PDZ domain-mediated interaction of interleukin-16 precursor proteins with myosin phosphatase targeting subunits
PDZ domain-mediated interaction of interleukin-16 precursor proteins with myosin phosphatase targeting subunits
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DOI：
10.1074/jbc.m306669200
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发表时间：
2003-10-24
影响因子：
4.8
通讯作者：
Kurth, R
中科院分区：
文献类型：
--
作者：
Bannert, N;Vollhardt, K;Kurth, R
细胞因子白介素-16是由caspase-3的两个大前体亚型的转录后切割产生的。较小的67 kDa蛋白（pro-IL-16）在免疫系统细胞中表达，包含三个PDZ（突触后密度/椎间盘大/区闭塞-1）结构域，而较大的141-kDa神经元变体（npro-IL-16）在其n端延伸处有两个额外的PDZ结构域，与神经元离子通道相互作用。使用酵母双杂交方法，我们已经确定了三个密切相关的肌球蛋白磷酸酶靶向亚基，MYPT1， MYPT2和MBS85，作为IL-16前体蛋白的结合伙伴。这些相互作用通过下拉测定、共免疫沉淀和等离子体共振实验得到验证。结合需要pro-IL-16完整的PDZ2结构域和配体中高度相关的c端区域，包括短亮氨酸拉链和-1位置不可或缺的丝氨酸，这提示了一种新颖的非常规PDZ结合模式。在转染的COS-7细胞中，亲il -16和肌球蛋白磷酸酶靶向亚基沿肌动球蛋白丝和应激纤维共定位。MYPT1、MYPT2和MBS85通过调节和靶向催化磷酸酶亚基到底物，调节肌肉和非肌肉细胞的各种收缩过程。我们的研究结果表明，IL-16前体分子参与了基于肌球蛋白的收缩过程，最有可能是在细胞运动中，为成熟细胞因子的趋化活性提供了功能联系。另外，神经元中由npro-IL-16、离子通道和肌球蛋白马达组成的细胞内复合体可能在蛋白质靶向中起作用。
The cytokine interleukin-16 is generated by posttranscriptional cleavage by caspase-3 of two large precursor isoforms. The smaller protein of 67 kDa (pro-IL-16) is expressed in cells of the immune system and contains three PDZ (postsynaptic density/disc large/zona occludens-1) domains, whereas the larger 141-kDa neuronal variant (npro-IL-16) has two additional PDZ domains in its N-terminal extension that interact with neuronal ion channels. Using the yeast two-hybrid approach we have identified three closely related myosin phosphatase targeting subunits, MYPT1, MYPT2, and MBS85, as binding partners of the IL-16 precursor proteins. These interactions were verified using pull-down assays, coimmunoprecipitations, and plasmon resonance experiments. Binding requires the intact PDZ2 domain of pro-IL-16 and highly related C-terminal regions in the ligands consisting of a short leucine zipper and an indispensable serine at the -1 position, suggesting a novel unconventional PDZ binding mode. Pro-IL-16 and the myosin phosphatase targeting subunits colocalize along actomyosin filaments and stress fibers in transfected COS-7 cells. By modulating and targeting the catalytic phosphatase subunit to its substrates, MYPT1, MYPT2, and MBS85 regulate various contractile processes in muscle and non-muscle cells. Our findings indicate an involvement of the IL-16 precursor molecules in myosin-based contractile processes, most likely in cell motility, providing a functional link to the chemotactic activity of the mature cytokine. Alternatively, an intracellular complex of npro-IL-16, ion channels, and components of myosin motors in neurons suggests a role in protein targeting.