Ⅰ.脂毒性应激对哺乳动物细胞周期的影响 脂毒性被认为是肥胖诱发糖尿病等代谢性疾病的机理之一。棕榈酸是人体血浆中含量最高的一种长链饱和脂肪酸。长时间高浓度的棕榈酸刺激可导致机体胰岛素抵抗，并诱发多种类型细胞(如胰岛β细胞等)的凋亡。因此，高浓度的棕榈酸可以用来模拟脂毒性环境以研究脂毒性应激对机体细胞造成的危害及其相关机理，从而为通过抵抗脂毒性治疗肥胖、糖尿病等代谢性疾病提供依据。 目前人们对脂毒性环境下细胞增殖的情况还知之甚少，本文的研究集中探讨脂毒性应激对细胞周期的影响及其分子机理。我们用高浓度的棕榈酸处理HeLa和C2C12等常见的哺乳动物细胞系，从而研究脂毒性应激对细胞周期进程可能的影响。流式细胞分析的结果显示，棕榈酸可导致显著的G2/M期阻滞。重要的是，我们发现棕榈酸通过抑制胞质分裂过程导致了双核细胞的积累。进一步的研究表明：在棕榈酸处理后，调控胞质分裂的关键因子小G蛋白RhoA活性受到明显抑制，同时对胞质分裂至关重要的微管骨架也出现紊乱。另外，我们发现蛋白质的棕榈酰化修饰抑制剂和一种常见的不饱和脂肪酸油酸均可抑制由棕榈酸引起的双核细胞的积累。 我们的研究结果揭示了一种新的脂毒性现象，即高浓度的棕榈酸抑制细胞的胞质分裂过程。这为我们进一步探索在脂毒性环境中体内处于活跃增殖状态的细胞，如造血干细胞、生殖细胞在分裂过程中可能出现的异常提供了重要线索。同时我们的结果也提示棕榈酸造成的胞质分裂异常可能涉及某些重要蛋白质的过度棕榈酰化修饰。因此，在脂毒性条件下蛋白质的过度棕榈酰化可能是造成细胞毒性的原因之一，蛋白质的棕榈酰化修饰抑制剂有可能成为抵抗脂毒性的潜在药物。 Ⅱ.核仁蛋白B23与PES1的相互作用及其生物学功能探讨 斑马鱼Pescadillo蛋白在人中的同源蛋白PES1是近年来新发现的一个重要的核仁蛋白，与核糖体生物发生过程及细胞周期调控密切相关。我们通过免疫共沉淀和免疫荧光的方法研究发现，核仁的标志蛋白之一B23蛋白与PES1蛋白存在相互作用及细胞内共定位。此外，我们用RNAi技术把B23蛋白下调后发现PES1蛋白出现了明显的核质积累。这些结果提示PES1蛋白有可能通过与B23蛋白的相互作用而定位于核仁。为了验证这一结论，我们进一步用Pull-down的办法鉴定出二者相互作用的区段。与我们的结论一致是，丧失与B23蛋白结合能力的PES1突变体不能定位于核仁。该部分结果支持了过去人们提出的B23蛋白作为一个核仁枢纽蛋白的假说，也使我们对PES1蛋白的核仁定位机制有了更深入的认识。 关键词：棕榈酸，脂毒性，棕榈酰化，细胞周期，胞质分裂，B23，PES1，核仁

 Ⅰ.The Effects of Lipotoxic Stress on Cell Cycle of Mammalian Cells Lipotoxicity has been implicated in the pathogenesis of obesity-associated diabetes and other metabolic diseases. Palmitate is the most abundant saturated free fatty acid in human plasma. It has been shown that a long-term exposure of the body to a high concentration of palmitate results in insulin resistance and induces apoptosis of many cell types such as pancreatic β cells. Thus, a high concentration of palmitate has been widely used as a lipotoxic stress model to study lipotoxicity and its related biological mechanisms. Currently little is known about how cells proliferate in a lipotoxic environment. We investigated the effects of lipotoxic stress on cell cycle progression by treating commonly-used mammalian cell lines such as HeLa and C2C12 cells with a high concentration of palmitate. Flow cytometry analysis showed that palmitate caused a dramatic G2/M cell-cycle arrest. We observed that palmitate induced binucleate cell formation via inhibition of cytokinesis. Further studies revealed that a small GTPase RhoA was inhibited upon palmitate treatment in association with disorganized microtubule cytoskeleton system. Moreover, we found that two palmitoylation inhibitors and the mono-unsaturated free fatty acid oleate could protect cells against palmitate-induced binucleate accumulation. Our results reveal a novel toxic effect of palmitate on cell division, suggesting that cytokinetic failure is a possible defection derived from lipotoxicity. Furthermore, our findings imply that excess palmitoylation of proteins may be involved in the palmitate-induced cytokinetic failure, and thereby palmitoylation inhibitors might be potential drugs against lipotoxicity. Ⅱ. The Interaction between B23 and PES1 and its Biological Implications PES1, the human homolog of zebrafish Pescadillo, is a nucleolar protein essential for ribosome biogenesis and cell proliferation. We identified B23, a nucleolar marker protein, as a novel interacting protein of PES1. Their interaction was verified by co-immunoprecipitation and immunofluorescence co-localization studies. Nucleoplasmic accumulation of PES1 was observed while B23 was knocked-down by RNAi. In addition, we mapped the interaction domains between PES1 and B23 via in vitro pull-down assays. Consistently, the PES1 mutant that lost binding activity to B23 was not able to locate at the nucleolus. Our findings support a hypothesis that B23 is a nucleolar hub protein for protein nucleolar targeting and provide an insight into the nucleolar localization mechanism of PES1. Key words: Palmitate, Lipotoxicity, Palmitoylation, Cell cycle, Cytokinesis, B23, PES1, Nucleolus