Endogenous Membrane Receptor Labeling by Reactive Cytokines and Growth Factors to Chase Their Dynamics in Live Cells
Endogenous Membrane Receptor Labeling by Reactive Cytokines and Growth Factors to Chase Their Dynamics in Live Cells
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DOI：
10.1016/j.chempr.2018.03.021
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发表时间：
2018-06-14
期刊：
影响因子：
23.5
通讯作者：
Hamachi, Itaru
中科院分区：
文献类型：
--
作者：
Takaoka, Yousuke;Uchinomiya, Shohei;Hamachi, Itaru
细胞因子和生长因子受体定位于细胞膜，在许多生物学事件中起着关键作用。由于受体的动态行为与细胞因子和生长因子依赖的信号通路密切相关，因此迫切需要可靠的技术对活细胞中的受体进行实时成像。在这里，我们描述了一种通过使用化学反应的细胞因子和生长因子与4-二甲氨基吡啶(DMAP)结合的方法来标记和成像活细胞中的膜受体。DMAP是一种促进酰基转移的有机催化剂。DMAP细胞因子和生长因子的构建依赖于低聚组氨酸标签和二聚体Ni(II)-三氮三乙酸酯之间的非共价原位配位化学。这种超分子方法相对简单和灵活，不仅可以用于标记瞬时表达的受体，也可以用于标记内源性表达的受体。此外，考虑到受体的这种无痕迹标记的好处，我们的技术可以实时成像活细胞中内源性表达的受体由配体诱导的动态。
Cytokine and growth factor receptors localized on cell membranes play key roles in many biological events. Because the dynamic behaviors of receptors are deeply involved in cytokine-and growth-factor-dependent signaling pathways, reliable techniques for real-time imaging of receptors in live cells are strongly desired. Here, we describe a method of covalently labeling and imaging membrane receptors in live cells by using chemically reactive cytokines and growth factors coupled with 4-dimethylaminopyridine (DMAP), an organo-catalyst that facilitates acyl transfer. Construction of the DMAP cytokines and growth factors relies on non-covalent coordination chemistry between an oligo-histidine tag and dimeric Ni(II)-nitrilotriacetate in an in situ manner. This supramolecular approach is relatively simple and flexible and can be applied to labeling not only transiently but also endogenously expressed receptors. Moreover, given the benefit of this traceless labeling of the receptors, our technique allows real-time imaging of the ligand-induced dynamics of endogenously expressed receptors in live cells.