Two novel cPLA2 binding proteins and cell death

cPLA 2与三种蛋白质相互作用，包括TipGO，我们称为PLIP的TipGO剪接变体和SIRT 2。本申请的目的是研究这些蛋白质在系膜细胞凋亡中的分子基础和作用。Tip 60和PLIP是增强细胞对凋亡的易感性的乙酰转移酶蛋白。我们最近证明PLIP表达与GO/G1期细胞的显著减少相关，而GO/G1期细胞与视网膜母细胞瘤蛋白的减少相关。肾小球系膜细胞内源性PLIP增加 提示PLIP可在DMA损伤后的G2/M检查点或之后诱导细胞凋亡。SIRT 2最近被鉴定为参与有丝分裂退出的调节蛋白。我们发现SIRT 2在遗传毒性应激后的系膜细胞中增加，其表达与TipGO的增加相关。我们发现，这些蛋白质在MRL/Faslpr小鼠的肾脏中减少，这表明在狼疮性肾小球肾炎的发展中具有潜在的作用。我们的假设是，PLIP，Tip 60和SIRT 2调节关键的细胞周期事件后，在系膜细胞中的DMA损伤，他们的功能是通过他们与cPLA 2的相互作用介导的，这些蛋白质的损失伴随和介导肾小球肾炎系膜的变化。本研究的主要目的是：1）研究Tip 60、PLIP和SIRT在正常系膜细胞周期中的表达及其cPLA 2依赖的功能; 2）研究TipGO和SIRT 2在DNA损伤诱导系膜细胞凋亡中的作用，以及pRb降解在PLIP相关凋亡中的作用; 3）确定TipGO细胞内运输差异的基础 和PLIP，并检查TipGO中潜在的核定位和核输出信号（内斯），并表征SIRT 2和TipGO剪接变体在MRL/Faslpr肾中的表达，以表明这些蛋白在肾小球肾炎发展中的潜在作用。我们将使用分子生物学和细胞生物化学的基本方法来回答这些问题。肾小球系膜细胞的细胞周期调控和细胞凋亡在肾小球肾炎的进展或消退中起着关键作用。更好地了解这些事件可能使我们能够开发出更好地治疗这些致残和昂贵疾病的疗法。

cPLA2 interacts with three proteins, including TipGO, a TipGO splice variant which we have called PLIP, and SIRT2. The goals of this application have been to study the molecular basis for and roles of these proteins in mesangial cell apoptosis. Tip60 and PLIP are acetyltransferase proteins that enhance cell susceptibility to apoptosis. We have recently demonstrated that PLIP expression is associated with a marked decrease in GO/G1 phase cells associated with a decrease in retinoblastoma protein. Endogenous PLIP is increased in mesangial cells following genotoxic stress suggesting that PLIP may induce apoptosis at or after the G2/M checkpoint following DMA damage. SIRT2 has recently been identified as a regulatory protein involved in mitotic exit. We show that SIRT2 is increased in mesangial cells following genotoxic stress and its expression is associated with an increase in TipGO. We show that these proteins are decreased in kidneys of MRL/Faslpr mice suggesting a potential role in the development of lupus glomerulonephritis. Our hypothesis is that PLIP, Tip60 and SIRT2 modulate critical cell cycle events following DMA damage in mesangial cells; that their functions are mediated via their interaction with cPLA2 and that loss of these proteins accompanies and mediates mesangial changes in glomerulonephritis. This proposal includes three specific Aims: 1) To determine the expression and cPLA2-dependent functions of Tip60, PLIP and SIRT during the normal cell cycle of mesangial cells; 2) To determine the roles of TipGO and SIRT2 in DNA-damage induced apoptosis in mesangial cells and to determine the role of pRb degradation in PLIP-associated apoptosis; 3) To determine the basis for differences in intracellular trafficking of TipGO and PLIP and examine potential nuclear localization and nuclear export signal (NES) in TipGO, and to characterize the expression of SIRT2 and TipGO splice variants in MRL/Faslpr kidneys in order to suggest a potential role for these proteins in the development of glomerulonephritis. We will use basic methods of molecular biology and cellular biochemistry to answer these questions. The regulation of cell cycle events and apoptosis in the mesangial cell is critical in the progression or resolution of glomerulonephritides. A better understanding of these events may allow us to develop therapies to better treat these incapacitating and costly diseases.