Cerbón Jorge, Falcon Alejandro, Hernández-Luna Carlos, Segura-Cobos David
Department of Biochemistry, Centro de Investigación y de Estudios Avanzados del IPN, 07000 México, D.F., Mexico.
Biochem J. 2005 May 15;388(Pt 1):169-76. doi: 10.1042/BJ20040475.
We recently reported that DAG (diacylglycerol) generated during sphingomyelin synthesis plays an important role in protein kinase C activation and cell proliferation in Madin-Darby canine kidney cells [Cerbon and Lopez-Sanchez (2003) Biochem. J. 373, 917-924]. In yeast cells, IPC (inositol phosphoceramide) synthase catalyses the transfer of phosphoinositol from phosphatidylinositol to ceramide to form IPC and generates DAG. In the present study, we found that, during the G1 to S transition after N2-starvation, there was a significant increase in the synthesis of IPC accompanied by a progressive increase (up to 6-fold) in the level of DAG. The increased DAG levels coincided with decrements in ceramide and sphingoid base levels, conditions that are adequate for the activation of putative protein kinase C required for the G1 to S transition and proliferation of yeast cells. To separate the role of DAG generated during IPC synthesis from that originating from other sources, we utilized beta-chloroalanine and myriocin, inhibitors of serine:palmitoyl-CoA transferase, the first committed step in sphingolipid synthesis, to avoid accumulation of sphingolipid intermediates. When the synthesis of sphingolipids was inhibited, DAG accumulation was significantly decreased and the G1 to S transition was blocked; such blockage was avoided by metabolic complementation with phytosphingosine. The DAG/ceramide ratio was 0.27 and it changed to 2.0 during growth re-initiation, suggesting that the synthesis of phosphosphingolipids could act to switch growth arrest (increased ceramide) to a mitogenic signal (increased DAG), and that this signalling process is preserved in yeast and mammalian cells.
我们最近报道，在鞘磷脂合成过程中产生的二酰基甘油（DAG）在蛋白激酶C激活和Madin-Darby犬肾细胞的细胞增殖中起重要作用[Cerbon和Lopez-Sanchez（2003年）《生物化学杂志》373卷，917 - 924页]。在酵母细胞中，肌醇磷酸神经酰胺（IPC）合酶催化磷酸肌醇从磷脂酰肌醇转移至神经酰胺以形成IPC，并产生DAG。在本研究中，我们发现，在N2饥饿后的G1期至S期转变过程中，IPC的合成显著增加，同时DAG水平逐渐升高（高达6倍）。DAG水平的升高与神经酰胺和鞘氨醇碱水平的降低同时出现，这些条件足以激活酵母细胞从G1期至S期转变及增殖所需的假定蛋白激酶C。为了区分IPC合成过程中产生的DAG与其他来源产生的DAG的作用，我们使用了β-氯丙氨酸和myriocin，它们是丝氨酸：棕榈酰辅酶A转移酶的抑制剂，该酶是鞘脂合成的第一个关键步骤，以避免鞘脂中间体的积累。当鞘脂合成受到抑制时，DAG积累显著减少，G1期至S期转变被阻断；通过植烷酸的代谢互补可避免这种阻断。DAG/神经酰胺的比率为0.27，在重新开始生长时变为2.0，这表明磷酸鞘脂的合成可以将生长停滞（神经酰胺增加）转变为促有丝分裂信号（DAG增加），并且这种信号传导过程在酵母和哺乳动物细胞中都得以保留。
