CELL GROWTH REGULATION VIA THE MEKK/SAPK CASCADE

我们以前的特点是MEKK 1蛋白激酶作为一个上游介质的应激激活蛋白激酶级联。应激激活蛋白激酶（SAPK，又称JNK）级联反应由于参与多种生理和病理刺激，特别是炎症细胞因子的调控，近年来受到了广泛的关注。同样重要的是MEKK 1在致癌作用中的作用尚未阐明。 应激激活级联反应受DNA损伤剂刺激，导致程序性细胞死亡（凋亡），常用于化疗策略。MEKK 1和SAPK被广泛认为在凋亡反应中起作用，尽管我们最近的数据表明MEKK 1对凋亡的影响是复杂的，并且对细胞死亡可以是积极的或消极的。 通过MEKK 1进行信号传导的中心问题是1）其活化方式，以及2）其将“应激信号”传递到细胞生长机制的机制。MEKK 1是一个200 kD的蛋白质，具有一个大的氨基末端调节结构域，具有未知的功能。 虽然MEKK 1在催化结构域内被磷酸化，但我们最近的研究结果表明磷酸化不是MEKK 1功能调节的主要机制。相反，我们最近获得的证据表明，两个非常不寻常的机制调节MEKK 1功能。 其中第一个是MEKK 1的靶向蛋白水解。我们已经确定了三个不同的蛋白水解酶切位点MEKK 1，并假设不同的刺激事件激活MEKK 1在功能上可区分的方式。 在细胞凋亡过程中，称为半胱天冬酶的蛋白酶切割MEKK 1并将其活性从抗细胞凋亡NF κ B途径重定向，而是促进SAPK介导的AP 1转录因子途径的激活。 其次，我们已经确定了活性醌作为有效的激活剂或抑制剂的SAPK途径（取决于浓度）和两个电子醌还原酶NQO 1作为关键的压力信号传导途径的MEKK 1上游的传输。 我们目前的数据导致的结论，巯基氧化还原状态的调节是MEKK 1激活的主要手段，并假设MEKK 1本身是通过其氨基末端的氧化还原调节的目标。考虑到应激信号的下游功能，我们发现SAPK蛋白激酶在细胞周期转换期间，即在G2/M期间被激活。 此外，MEKK 1的表达或用各种应激剂处理细胞导致与细胞周期蛋白B相关的CDK激酶活性的抑制。 我们建议的特点，应激信号与细胞周期蛋白B/cdc 2激酶相互作用的机制。

We have previously characterized the MEKK1 protein kinase as an upstream mediator of the Stress Activated Protein Kinase cascade. Stress Activated Protein Kinase (SAPK, alternatively known as JNK) cascade has recently been closely investigated owing to its participation in many physiological and pathologic stimuli, particularly inflammatory cytokines. Also of central importance is the yet unclarified role of MEKK1 in carcinogenesis. The Stress-Activated cascade is stimulated by DNA damaging agents that lead to programmed cell death (apoptosis) and are commonly used in chemotherapeutic strategies. MEKK1 and SAPK are broadly assumed to play a role in the apoptotic response, though our recent data suggest that effects of MEKK1 on apoptosis are complex and can be either positive or negative towards cell death. Central questions in signaling through MEKK1 are 1) its means of activation, and 2) its mechanism of transmitting the "stress signal" through to cell growth machinery. MEKK1 is a 200 kD protein with a large amino terminal regulatory domain that has uncharacterized function. While MEKK1 is phosphorylated within the catalytic domain, our recent results indicate that phosphorylation is not the major mechanism of regulation of MEKK1 function. We have instead obtained recent evidence that two highly unusual mechanisms regulate MEKK1 functions. The first of these is targeted proteolysis of MEKK1. We have identified three distinct proteolytic cleavage sites on MEKK1, and hypothesize that different stimulatory events activate MEKK1 in functionally distinguishable ways. During apoptosis, proteases termed caspases cleave MEKK1 and redirect its activity away from the anti-apoptotic NFkappaB pathway, and instead promote activation of the SAPK-mediated AP1 transcription factor pathway. Secondly, we have identified reactive quinones as potent activators or inhibitors of the SAPK pathway (depending on concentration) and the two electron quinone reductase NQO1 as critical for transmission of stress signaling pathways upstream of MEKK1. We present data leading to the conclusion that regulation of sulfhydryl redox status is a prime means of MEKK1 activation, and hypothesize that MEKK1 itself is a target for redox regulation via its amino terminus. Considering the downstream function of stress signaling, we have found that the SAPK protein kinase becomes activated during cell cycle transition, during G2/M. Additionally, expression of MEKK1 or treatment of cells with various stress agents leads to inhibition of CDK kinase activity associated with cyclin B. We propose to characterize the mechanism by which stress signals interact with the cyclin B/cdc2 kinase.