Department of Molecular Therapeutics, and Translational Research Institute, The Scripps Research Institute, 130 Scripps Way, Jupiter, Florida 33458, United States.
ACS Chem Biol. 2011 Aug 19;6(8):808-18. doi: 10.1021/cb200062a. Epub 2011 May 24.
The c-jun N-terminal kinases (JNKs) are responsive to stress stimuli leading to activation of proapoptotic proteins and transcription. Additionally, JNK mitochondrial localization has been reported. To selectively target mitochondrial JNK signaling, we exploited JNK interaction with its mitochondrial scaffold, Sab, using small interfering RNAs (siRNAs) and a cell-permeable peptide corresponding to the KIM1 domain of Sab. Gene silencing and peptide interference of this interaction disrupted JNK translocation to the mitochondria and reduced phosphorylation of Bcl-2 without significant impact on c-Jun phosphorylation or AP-1 transcription. In contrast, the JNK inhibitory peptide (TI-JIP1) prevented these three functions. Tat-Sab(KIM1) selectivity was also demonstrated in anisomycin-stressed HeLa cells where Tat-Sab(KIM1) prevented Bcl-2 phosphorylation, cell death, loss of mitochondrial membrane potential, and superoxide generation but not c-Jun phosphorylation. Conversely, TI-JIP1 prevented all aforementioned stress-induced events. This probe introduces a means to evaluate JNK-mediated events on the mitochondria without intervening in nuclear functions of JNK.
c-jun N 端激酶（JNKs）对应激刺激有反应，导致促凋亡蛋白和转录的激活。此外，已经报道了 JNK 的线粒体定位。为了选择性地靶向线粒体 JNK 信号，我们利用 JNK 与其线粒体支架 Sab 的相互作用，使用小干扰 RNA（siRNA）和对应于 Sab 的 KIM1 结构域的细胞渗透性肽。该相互作用的基因沉默和肽干扰破坏了 JNK 向线粒体的易位，并减少了 Bcl-2 的磷酸化，而对 c-Jun 的磷酸化或 AP-1 转录没有显著影响。相比之下，JNK 抑制肽（TI-JIP1）阻止了这三种功能。在受到 anisomycin 应激的 HeLa 细胞中也证明了 Tat-Sab（KIM1）的选择性，其中 Tat-Sab（KIM1）阻止了 Bcl-2 的磷酸化、细胞死亡、线粒体膜电位的丧失和超氧化物的产生，但不阻止 c-Jun 的磷酸化。相反，TI-JIP1 阻止了所有上述应激诱导的事件。该探针引入了一种在不干预 JNK 核功能的情况下评估线粒体中 JNK 介导的事件的方法。
