MECHANISMS OF CHIEF CELL DEDIFFERENTIATION

项目摘要 我们研究损伤和炎症如何诱导成熟细胞，如消化酶分泌酶原的首席 细胞（ZC）分解其复杂的细胞结构并重新进入细胞周期。我们之前已经证明， ZC通过在许多组织中保守的一系列分子细胞事件而增殖， 在成熟细胞被招募回细胞周期以响应组织损伤的情况下， 因此，细胞有一个进化上保守的程序，这种重新编程，因为他们为死亡（凋亡） 分裂（mitosis）。我们称之为paligenosis，并表明成熟细胞：首先降解/回收 其分化的细胞组分（阶段1），然后诱导祖细胞样基因（例如，Sox 9 = 阶段2），最后重新进入细胞周期（阶段3）。Paligenotic ZCs转化为细胞， 在组织病理学上，作为在胃长期感染期间发生的化生类型， 幽门螺杆菌：假幽门或痉挛性多肽表达化生（SPEM）。 化生可以随着组织修复而消退，或者变成慢性并增加进展为 发育不良和癌症。我们已经表明，黄褐斑是由mTORC 1的动态变化， 细胞翻译控制蛋白复合物。在ZC中，mTORC 1在基线时升高，以驱动 消化酶，它在第一阶段关闭，并在第三阶段重新激活。如果没有mTORC 1， 第2阶段，细胞看起来化生，但无法进入S期。 在这里，我们探讨的机制，诱导和促进paligenosis。我们展示了初步数据 暗示综合应激反应（ISR）途径作为一个中心paligenosis枢纽，具有特殊的作用， 与ISR相关的转录因子Atf 3，以及另一个我们假设是一个 ATF 3的靶点：Ifrd 1，一种多功能支架蛋白。我们假设大规模组织的应力 损伤和/或炎症触发ISR活动过度，导致ATF 3和IFRD 1大大增加， 帮助细胞回到细胞周期。在缺乏Atf 3或Ifrd 1的情况下，我们表明Paligenosis是有缺陷的。我们 具体目标将是：1）确认并进一步表征ISR处于哪个阶段，并确认和 部分使用Atf 3 −/−小鼠表征ATF 3的作用; 2）鉴定ISR中涉及的其他基因， 通过用Ifrd 1 −/−小鼠证实IFRD 1的作用，探测ATF 3和IFRD 1的相对贡献， 通过对双基因敲除小鼠的特征分析，最后在paligenosis ± ATF 3中进行ChIP-和RNA-Seq; 3)为了测试已知的ATF 3和ISR基因以及Aim 2中开发的新靶标， 疾病模型（例如幽门螺杆菌慢性感染小鼠）、人翻译样品（组织微阵列 以及另外的近1000名患者的化生和癌症的人类样本），以及在小鼠模型中， 肿瘤发生± ATF 3。总之，这些实验可以帮助我们了解细胞使用的基本机制 在再生和肿瘤发生中的作用，这不仅适用于胃，也可能适用于其他器官。

PROJECT SUMMARY We study how injury and inflammation induce mature cells like the digestive-enzyme-secreting zymogenic chief cell (ZC) to disassemble their complex cell architecture and re-enter the cell cycle. We previously showed that ZCs become proliferative via a sequence of molecular-cellular events conserved across many tissues and species in scenarios where mature cells are recruited back into the cell cycle in response to tissue damage. Thus, cells have an evolutionarily conserved program for this reprogramming, as they do for death (apoptosis) and division (mitosis). We call this program paligenosis and showed that mature cells: first degrade/recycle their differentiated cell components (Stage 1), then induce expression of progenitor-like genes (eg. Sox9 = Stage 2), and finally re-enter the cell cycle (Stage 3). Paligenotic ZCs convert to cells that can be seen histopathologically as the type of metaplasia that occurs in stomach during long-term infection with the bacterium Helicobacter pylori: pseudopyloric or Spasmolytic Polypeptide Expressing Metaplasia (SPEM). Metaplasia can either resolve as tissue is repaired or become chronic and increase risk for progression to dysplasia and cancer. We have shown that paligenosis is governed by dynamic changes in mTORC1, the cellular translation control protein complex. mTORC1 is elevated at baseline in ZCs to drive translation of digestive enzymes, it shuts off at Stage 1, and reactivates at Stage 3. Without mTORC1, paligenosis stops at Stage 2 with cells looking metaplastic, but unable to enter S-phase. Here, we explore the mechanisms that induce and promote paligenosis. We show preliminary data implicating the Integrated Stress Response (ISR) pathway as a central paligenosis hub with a particular role for the transcription factor Atf3, which is associated with the ISR, and another gene which we hypothesize is a target of ATF3: Ifrd1, a multifunctional scaffolding protein. We hypothesize that the stress of large-scale tissue damage and/or inflammation triggers ISR hyperactivity, which leads to greatly increased ATF3 and IFRD1, to help push cells back into the cell cycle. In the absence of Atf3 or Ifrd1, we show paligenosis is defective. Our Specific Aims will be: 1) to confirm and further characterize at which stages ISR is active and confirm and characterize the role for ATF3 using, in part Atf3−/− mice; 2) to identify additional genes involved in the ISR and paligenosis by confirming the role of IFRD1 with Ifrd1−/− mice, probe relative contributions of ATF3 and IFRD1 by characterizing double knockout mice, and finally to perform ChIP- and RNA- Seq during paligenosis ±ATF3; 3) to test known ATF3 and ISR genes and new targets developed in Aim 2 in a pipeline of more physiological disease models (eg chronic infection of mice with H pylori), human translational samples (Tissue Microarray and additional human samples of metaplasia and cancer with nearly a 1000 patients), and in a mouse model of tumorigenesis ±ATF3. Together, the experiments may help us understand fundamental mechanisms cells use in regeneration and tumorigenesis that apply not just to the stomach but potentially other organs as well.