Dynamic positive feedback phosphorylation of mixed lineage kinase 3 by JNK reversibly regulates its distribution to triton-soluble domains
Dynamic positive feedback phosphorylation of mixed lineage kinase 3 by JNK reversibly regulates its distribution to triton-soluble domains
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DOI：
10.1074/jbc.m603324200
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发表时间：
2006-07-14
影响因子：
4.8
通讯作者：
Gallo, Kathleen A.
中科院分区：
文献类型：
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作者：
Schachter, Karen A.;Du, Yan;Gallo, Kathleen A.
MLK3(Mix Lineage Kinase 3)是一种广泛表达的哺乳动物丝氨酸/苏氨酸蛋白激酶，可激活多种MAPK通路。在此之前，我们实验室使用活体标记/质谱法来鉴定活化的MLK3的磷酸化位点。在11个已鉴定的位点中，有7个对应于由脯氨酸导向的激酶进行磷酸化的共识基序。基于这些结果，我们假设JNK，或另一种脯氨酸导向的激酶，作为反馈环的一部分，使MLK3磷酸化。在此，我们提供了JNK在体外和体内都能使MLK3磷酸化的证据。阻断JNK导致MLK3去磷酸化。MLK3的低磷酸化形式是不活跃的，并重新分布为不溶于Triton的部分。从JNK抑制中恢复可以恢复MLK3的溶解度和活性，表明重新分配过程是可逆的。本工作描述了一种新的MLK3调控模式，JNK介导的MLK3反馈磷酸化通过在Triton可溶形式和Triton不可溶形式之间循环来调节MLK3的激活和失活状态。
MLK3 (mixed lineage kinase 3) is a widely expressed, mammalian serine/threonine protein kinase that activates multiple MAPK pathways. Previously our laboratory used in vivo labeling/mass spectrometry to identify phosphorylation sites of activated MLK3. Seven of 11 identified sites correspond to the consensus motif for phosphorylation by proline-directed kinases. Based on these results, we hypothesized that JNK, or another proline-directed kinase, phosphorylates MLK3 as part of a feedback loop. Herein we provide evidence that MLK3 can be phosphorylated by JNK in vitro and in vivo. Blockade of JNK results in dephosphorylation of MLK3. The hypophosphorylated form of MLK3 is inactive and redistributes to a Triton-insoluble fraction. Recovery from JNK inhibition restores MLK3 solubility and activity, indicating that the redistribution process is reversible. This work describes a novel mode of regulation of MLK3, by which JNK-mediated feedback phosphorylation of MLK3 regulates its activation and deactivation states by cycling between Triton-soluble and Triton-insoluble forms.