Control of genetic and epigenetic instabilities by lincRNA genes

 描述(由申请人提供)：癌细胞在许多重要特征上不同于正常细胞。癌症的这些“特征”影响许多生物过程，并且是在人类癌症的多步骤发展过程中获得的。毫不奇怪，遗传和/或表观遗传变化存在于大多数癌症中，并被认为是导致这些表型变化的原因。此外，遗传和/或表观遗传不稳定性被认为存在于大多数癌症中，并被认为是产生个体肿瘤细胞中存在的大量遗传和/或表观遗传变化所必需的。了解推动这些变化的机制显然是一个重要的目标。我的实验室之前描述了一种与某些肿瘤衍生的染色体重排相关的染色体异常。这种异常表型影响个体染色体，以延迟复制时间(DRT)、延迟有丝分裂染色体凝集(DMC)和频繁的染色体重排为特征。我们开发了一种‘染色体工程’系统，使我们能够以高效和可重复的方式产生DRT/DMC染色体。因此，使用基于Cre/loxP的策略来筛选人类染色体中的重排，我们发现了5个平衡易位，影响了8个不同的常染色体，都表现为DRT/DMC。我们还发现，工程的DRT/DMC染色体表现出染色体结构的不稳定性。最近，我们发现Cre/loxP介导的一个长的非编码RNA基因，我们称之为6号染色体上的异步复制和常染色体RNA(ASAR6)的破坏，导致了人类6号染色体上的DRT/DMC。我们还发现，ASAR6与Xist非编码RNA基因有许多共同的特征，包括：随机的单等位基因表达，异步复制，顺式基因中其他单等位基因的沉默，以及在基因组转基因异位整合后延迟整个染色体的复制。值得注意的是，Xist的缺失会导致X染色体复制延迟、染色质结构异常和次级重排。因此，ASAR6的破坏导致Xist缺失的表型复制。此外，Xist的破坏最近被证明在小鼠中导致100%穿透性血液系统恶性肿瘤，表明Xist作为肿瘤抑制因子发挥作用。最近，我们发现了第二个非编码RNA基因，暂定为ASAR15，它控制着人类15号染色体的复制时机和稳定性。综上所述，这些观察结果提出了一种有趣的可能性，即所有哺乳动物的染色体都含有类似Xist的肿瘤抑制基因，其功能是维持单个染色体的遗传和表观遗传稳定性。组成这一建议的实验旨在：1)表征与ASAR6、ASAR15或XIST中断相关的结构重排的“分子特征”；2)阐明与ASAR6或ASAR15中断相关的染色质和基因表达变化；以及3)识别具有ASAR6、ASAR15或XIST结构重排的人类癌细胞。

 DESCRIPTION (provided by applicant): Cancer cells differ from their normal cellular counterparts in many important characteristics. These `hallmarks' of cancer affect numerous biological processes, and are acquired during the multistep development of human cancer. Not surprisingly, genetic and/or epigenetic changes are present in most if not all cancers, and are known to be responsible for these phenotypic alterations. In addition, genetic and/or epigenetic instabilities are thought to be present in most cancers and are thought to be required to generate the numerous genetic and/or epigenetic changes that are present in individual tumor cells. Understanding the mechanisms that drive these alterations is clearly an important goal. My lab previously characterized a chromosomal abnormality associated with certain tumor-derived chromosome rearrangements. This abnormal phenotype affects individual chromosomes and is characterized by delayed replication timing (DRT), delayed mitotic chromosome condensation (DMC), and frequent rearrangement of the affected chromosomes. We have developed a `chromosome engineering' system that allows us to generate DRT/DMC chromosomes in an efficient and reproducible manner. Thus, using a Cre/loxP-based strategy to screen for rearrangements in human chromosomes we identified 5 balanced translocations, affecting 8 different autosomes, all displaying DRT/DMC. We also found that the engineered DRT/DMC chromosomes display chromosome structure instability. More recently, we found that Cre/loxP-mediated disruption of a long non-coding RNA gene, which we named ASynchronous replication and Autosomal RNA on chromosome 6 (ASAR6), results in DRT/DMC on human chromosome 6. We also found that ASAR6 shares many characteristics with the Xist non-coding RNA gene, including: random monoallelic expression, asynchronous replication, silencing of other monoallelic genes in cis, and the ability to delay replication of entire chromosomes following ectopic integration of genomic transgenes. Notably, deletion of Xist results in delayed replication, abnormal chromatin structure and secondary rearrangements on the X chromosome. Therefore, disruption of ASAR6 results in a phenocopy of Xist deletion. Furthermore, disruption of Xist was recently shown to cause a 100% penetrant hematopoetic malignancy in mice, indicating that Xist functions as a tumor suppressor. More recently, we identified a second non-coding RNA gene, tentatively named ASAR15, which controls replication timing and stability of human chromosome 15. Taken together, these observations raise the intriguing possibility that all mammalian chromosomes contain `Xist-like' tumor suppressor loci functioning to maintain the genetic and epigenetic stability of individual chromosomes. The experiments that make up this proposal are designed to: 1) characterize the "molecular signature" of structural rearrangements associated with ASAR6, ASAR15 or Xist disruption; 2) elucidate the chromatin and gene expression changes associated with ASAR6 or ASAR15 disruption; and 3) identify human cancer cells with structural rearrangements in ASAR6, ASAR15 or XIST.