MECHANISMS OF CHIEF CELL DEDIFFERENTIATION

 描述(申请人提供)：仅在过去的两年里，消化器官中的许多成熟细胞系就可以去分化并重新进入细胞周期，以修复组织损伤。我们和其他人已经证明，有丝分裂后的细胞可以重新诱导祖细胞标志物，重新进入细胞周期，甚至可以作为其他血统的祖细胞。然而，去分化过程是有风险的，因为重新表达祖细胞基因是细胞化生的一种形式。在慢性损伤的状态下，比如环胃的幽门螺杆菌感染，化生无限期地持续下去，并增加进展为癌症的风险。总体而言，细胞去分化为理解组织修复、“干性”和癌症的发生提供了一个新的范式。我们的首要目标是了解控制去分化的细胞和分子机制。在这里，我们将集中在胃消化-分泌酶原主细胞(ZCS)去分化的早期阶段。我们的初步和已发表的数据将表明，第一阶段是溶酶体吞噬分泌颗粒，消化酶分泌颗粒(分泌器官“缩小”)的溶酶体周转必须发生在进入下一个阶段之前：前体细胞基因的化生重新表达和细胞周期重新进入。我们将发现，去分化ZCS最早的分子变化是bHLH转录因子MIST1(BHLHA15)的表达显著减少，即使在健康的ZCS中，Mist1的缺失也足以诱导分泌颗粒的溶酶体翻转。最后，我们将证明，当MIST1在去分化过程中丢失时，MIST1唯一已知的运输溶酶体的转录目标是RAB26，这是溶酶体攻击分泌颗粒的最有可能的原因。我们的目标是：1)确定MIST1在化生下调中的必要性/充分性；2)确定RAB26是否通过自噬/mTOR信号影响通量，以及它是否调节溶酶体和/或自溶酶体对颗粒的破坏；3)确定在成熟的ZCS中关键的自噬基因ATG5和ATG7缺失后，是否需要自噬机制来诱导化生；b)在有ZCS的去分化/化生病灶的标本数据库中，定量自噬/溶酶体与分泌颗粒的相互作用。这些实验将首次揭示新描述的细胞去分化过程背后的分子机制，并将对人类健康产生影响，因为我们需要了解成熟细胞的去分化是如何形成化生前体病变的，以便能够了解如何逆转它们，降低患者进展为癌症的风险。

 DESCRIPTION (provided by applicant): It has become clear in just the past two years that many mature cell lineages in digestive organs can dedifferentiate and re-enter the cell cycle to repair tissue damage. We and others have shown that post-mitotic cells can re-induce progenitor cell markers, re-enter the cell cycle, and can even serve as progenitors for other lineages. The dedifferentiation process carries a risk, however, as re-expressing progenitor cell genes is a form of cellular metaplasia. In states of chronic injury, like H. pylori infection in huan stomach, metaplasia continues indefinitely and increases risk for progression to cancer. Overall, cellular dedifferentiation presents a new paradigm for understanding tissue repair, "stemness", and cancer initiation. Our overarching goal is to understand the cellular and molecular mechanisms governing dedifferentiation. Here, we will focus on the earliest stages of dedifferentiation in gastric digestive-enzyme secreting zymogenic chief cells (ZCs). Our preliminary and published data will show that the first stage is engulfment of secretory granules by lysosomes and that lysosomal turnover of the digestive enzyme secretory granules (secretory apparatus "downscaling") must occur before progression to the next stages: metaplastic re- expression of progenitor cell genes and cell cycle re-entry. We will show the earliest molecular change in dedifferentiating ZCs is dramatically decreased expression of the bHLH transcription factor MIST1 (BHLHA15) and that deletion of Mist1 even in healthy ZCs is sufficient to induce lysosomal turnover of secretory granules. Finally, we will show that the only known transcriptional target of MIST1 that traffics lysosomes and is hence the most likely reason for the lysosomal attack on secretory granules when MIST1 is lost during dedifferentiation is the small GTPase, RAB26. Our aims are to: 1) determine the requirement/sufficiency for MIST1 in metaplasia downscaling; 2) determine if RAB26 affects flux through autophagy/mTOR signaling and whether it modulates granule destruction by lysosomes and/or autolysosomes; 3) a) determine if autophagic machinery is required for downscaling by inducing metaplasia following deletion in mature ZCs of the key autophagy genes Atg5 and Atg7 and b) to quantify autophagy/lysosome interaction with secretory granules in a database of human specimens where there are foci of ZCs undergoing dedifferentiation/metaplasia. Together the experiments will uncover for the first time the molecular mechanisms underlying the newly described cellular process of dedifferentiation and will have impact on human health because we will need to understand how metaplastic precursor lesions form from dedifferentiation of mature cells to be able to understand how to revert them and decrease risk for patients to progress to cancer.