Discovery and analysis of dovel lipid signal transduction complexes

二酰基甘油（DAG）激酶（DGK）通过磷酸化DAG以产生PA来调节两种信号脂质DAG和磷脂酸（PA）之间的平衡。到目前为止，已经鉴定了10种哺乳动物DGK同工酶。最近的研究发现DGK同工酶在多种信号转导途径中起着重要作用，我们发现DGK α在几种人黑色素瘤细胞系中表达，但在非癌黑色素细胞中不表达。有趣的是，野生型DGKα的过度表达，而不是其激酶死亡突变体，显著抑制了肿瘤坏死因子-a诱导的阿基人黑素瘤细胞凋亡。在反向实验中，小干扰RNA介导的DGKα敲低显著增强了细胞凋亡，从而令人信服地表明该异构体具有细胞保护功能。此外，我们还获得了一些证据，表明DGKα通过激活细胞凋亡抑制肿瘤坏死因子-α诱导的黑色素瘤细胞凋亡。 ...更多信息 核因子κB离子。因此，该转录因子可能是DGKα下游的关键因子之一，在细胞增殖和存活过程中，我们已经证明DGKγ通过其催化作用作为Racl的上游抑制因子，从而形成板状伪足/膜皱褶。最近，我们进一步发现DGKγ特异性地与β2-chimaerin（一种Rac特异性GTP酶激活蛋白）相互作用并激活β2-chimaerin，响应于表皮生长因子和过氧化氢/佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）的细胞刺激。PMA被认为是主要需要的转换82-嵌合体的活性形式。另一方面，H_2O_2可诱导β2-chimaerin的Cl结构域释放Zn^<2+>。逐步缺失分析表明，β2-chimaerin的Src同源2（SH 2）和Cl结构域与DGKγ的N端半个催化区相互作用。总之，这些结果表明DGKγ和β2-嵌合蛋白（和Racl）之间的功能联系在响应广泛的细胞刺激方面具有广泛的意义。我们的工作提供了一种新的蛋白质-蛋白质相互作用机制，即SH 2结构域与Cl结构域协同作用的非磷酸酪氨酸依赖性相互作用。脂质磷酸磷酸酶（Lipid phosphate phosphatases，LPPs，type 2 phosphatidic acid phosphatases）是一种具有六个跨膜结构域的膜蛋白，可使多种细胞外脂质磷酸盐脱磷酸。虽然已知LPP 3与Triton X-100不溶性筏结合，但我们发现LPP 1也与脂筏（Triton X-100可溶，但CHAPS不溶性）相关，不同于那些含有LPP 3的脂筏。LPP 1和LPP 3分布在不同的脂筏中，这些脂筏可以提供限定其非冗余生理功能的独特微环境。少

Diacylglycerol (DAG) kinase (DGK) modulates the balance between the two signaling lipids, DAG and phosphatidic acid (PA), by phosphorylating DAG to yield PA. To date, ten mammalian DGK isozymes have been identified. Beyond our expectations, recent studies have revealed that DGK isozymes play pivotal roles in a wide variety of signal transduction pathways.Recently, we found that DGKa is expressed in several human melanoma cell lines but not in noncancerous melanocytes. Intriguingly, the overexpression of wild-type DGKα, but not of its kinase-dead mutant, markedly suppressed tumor necrosis factor-a-induced apoptosis of AKI human melanoma cells. In the reverse experiment, small interfering RNA-mediated knockdown of DGKα significantly enhanced the apoptosis, thus convincingly indicating that this isoform possesses a cytoprotective function. Moreover, we obtained several lines of evidence suggesting that DGKα suppresses tumor necrosis factor-a-induced melanoma cell apoptosis through activat … More ion of nuclear factor κB. Thus, this transcription factor is likely to be one of the key factors downstream of DGKα during cell proliferation and survival.We had already demonstrated that DGKγ functions through its catalytic action as an upstream suppressor of Racl, and consequently, lamellipodium/membrane ruffle formation. Recently, we further found that DGKγ specifically interacts with and activates β2-chimaerin, a Rac-specific GTPase-activating protein, in response to cell stimulation with epidermal growth factor and hydrogen peroxide/phorbol 12-myristate 13-acetate (PMA). PMA was found to be mainly required for a conversion of 82-chimaerin to an active form. On the other hand, H_2O_2 was suggested to induce a release of Zn^<2+> from the Cl domain of β2-chimaerin. By stepwise deletion analysis, we demonstrated that the Src homology 2 (SH2) and Cl domains of β2-chimaerin interacted with the N-terminal half of catalytic region of DGKγ. Taken together, these results suggest that the functional link between DGKγ and β2-chimaerin (and Racl) has a broad significance in response to a wide range of cell stimuli. Our work offers a novel mechanism of protein-protein interaction, that is, the phosphotyrosine-independent interaction of the SH2 domain acting in cooperation with the Cl domain.Lipid phosphate phosphatases (LPPs, type 2 phosphatidic acid phosphatases), integral membrane proteins with six transmembrane domains, dephosphorylate a variety of extracellular lipid phosphates. Although LPP3 is already known to bind to Triton X-100-insoluble rafts, we found that LPP1 is also associated with lipid rafts (Triton X-100-soluble but CHAPS-insoluble) distinct from those harboring LPP3. LPP1 and LPP3 are distributed in distinct lipid rafts that may provide unique microenvironments defining their non-redundant physiological functions. Less