Roles of the ER Stress Surveillance Pathway During the Cell Cycle

项目摘要 为了确保每个分裂的细胞接受一套完整的正确的基因组，细胞周期检查点在 在整个细胞周期中。然而，关于是否存在类似的检查站， 细胞质成分或细胞器。内质网（ER）是一个门户， 分泌途径，产生几乎所有的分泌和细胞表面膜蛋白，以及 合成细胞脂质以前，我们确定了一个细胞周期检查点，称为ER应激监测 （ERSU）途径，其确保在模式生物S中遗传足够水平的功能性ER。 啤酒。响应于ER应激，ERSU途径（1）阻断应激的ER遗传到 子细胞，（2）将隔蛋白环从芽颈（胞质分裂的部位）错误定位，并最终，（3） 导致胞质分裂时细胞周期暂时停滞，直到ER功能性稳态重新建立。的细胞 缺乏ERSU的组分，因此不能安装ERSU通路，在ER应激时死亡，强调 这个检查站的重要性。ERSU途径不同于研究充分的未折叠蛋白 反应我们发现，植物鞘氨醇（PHS），鞘脂的早期中间体， 生物合成，ER应激增加，启动ERSU标志事件。此外，我们定义了一个 PHS结合基序，其存在于reticulon家族蛋白的两个不同的跨膜结构域中（即， Rtn1和Yop1），导致ERSU事件的激活。在目前的提案中，在AIM 1中，我们将应用 分子和细胞生物学方法来剖析PHS如何与Rtn1或Yop1结合导致ERSU activation.在AIM 2中，我们将通过解剖ER脂毒性应激的影响来扩展ERSU的范围 诱导ER形态学改变，对ERSU分子事件。在AIM 3中，我们将研究ER如何 压力影响哺乳动物的细胞周期。在哺乳动物有丝分裂过程中， 在细胞中，功能性ER的分裂可能与核有丝分裂更紧密地编排在一起。 机制等我们对哺乳动物septin亚基的初步研究结果为以下蛋白的存在提供了很大的希望： 调控事件，可能共享相似的酵母ERSU。因此，我们将充分研究ER的影响 对（A）哺乳动物septin亚基和细胞动力学组分，以及（B）主要有丝分裂细胞周期的应激 涉及ER的结构变化，如ER从“有丝分裂禁区”和核内清除 拆卸和重新组装。了解整合ER稳态的分子机制 细胞周期事件将提供前所未有的洞察人类疾病所造成的失败ER 调控.

PROJECT SUMMARY To ensure that each dividing cell receives a complete set of the correct genome, cell cycle checkpoints are in place throughout the cell cycle. Yet, less is known about whether similar checkpoints exist for division of the cytoplasmic components or organelles of the cell. The endoplasmic reticulum (ER) is a gateway for the secretory pathway, generating almost all of the secreted and cell surface membrane proteins as well as synthesizing cellular lipids. Previously, we identified a cell cycle checkpoint, termed the ER stress surveillance (ERSU) pathway, that ensures the inheritance of sufficient levels of functional ER in the model organism S. cerevisiae. In response to ER stress, the ERSU pathway (1) blocks the inheritance of the stressed ER into the daughter cell, (2) mislocalizes the septin ring from the bud neck, the site of cytokinesis, and ultimately, (3) leads to temporary cell cycle arrest at cytokinesis until ER functional homeostasis is re-established. Cells that lack components of the ERSU, and thus cannot mount the ERSU pathway, die upon ER stress, underscoring the importance of this checkpoint. The ERSU pathway is distinct from the well-studied unfolded protein response. We have found that levels of phytosphingosine (PHS), an early intermediate of sphingolipid biosynthesis, increase upon ER stress, setting in motion the ERSU hallmark events. Moreover, we defined a PHS binding motif that is found within two different transmembrane domains of reticulon family proteins (i.e., Rtn1 and Yop1), leading to the activation of the ERSU events. In the current proposal, in AIM 1, we will apply molecular and cell biological approaches to dissect how PHS binding to Rtn1 or Yop1 results in ERSU activation. In AIM 2, we will extend the scope of the ERSU by dissecting the impact of ER lipotoxic stress induced by ER morphological changes, on the ERSU molecular events. In AIM 3, we will investigate how ER stress impacts the mammalian cell cycle. As the nuclear membrane breaks down during mitosis in mammalian cells, the division of functional ER may be even more tightly choreographed with the nuclear mitotic mechanisms. Our preliminary result of a mammalian septin subunit holds great promise for the presence of regulatory events that may share similarity to the yeast ERSU. Thus, we will fully investigate the impact of ER stress on (A) the mammalian septin subunits and cytokinetic components, and (B) major mitotic cell cycle structural changes that involve the ER, such as ER clearing from the “mitotic exclusion zone” and nuclear disassembly and reassembly. Understanding the molecular mechanisms that integrate ER homeostasis with cell cycle events will provide unprecedented insights into human diseases caused by the failure of ER regulation. .