RNF128 Regulation of TP53 in Barrett's Progression

摘要： 食管腺癌（EAC）是一种致命的癌症，发生在称为巴雷特的癌前病变区域 化生（BE）。EAC的发病率以高于任何其他癌症的速度增加， 患有胃食管反流病（GERD）和肥胖症。了解巴雷特的哪些特性 化生上皮与EAC进展相关，可能潜在地发现新的治疗方法 预防这种癌症的策略。我们利用从进展为高级别的患者中获得的BE， 异型增生和EAC，并对基因表达以及亚型特异性mRNA进行RNAseq分析 分析非发育异常的Barrett、低度发育异常（LGD）、高度发育异常（HGD）至EAC。 重要的是，我们观察到从Barrett/LGD到HGD/EAC的转变与显著的同种型相关， 个体基因的变化。这种“同种型转换”与途径分析一致，表明 剪接作为BE向EAC进展中改变的顶部途径。我们观察到，“同种型转换” 随着保护性粘蛋白的丢失，炎症增加，ATM/DNA损伤反应的激活， 在HGD和EAC中增加的H2 H2 AX染色。最近有报道称， 拼接我们的一个顶级基因，表现出戏剧性的同种型转换是RNF 128或圣杯，一种泛素 与野生型（WT）TP 53的N-末端功能性相互作用以靶向其降解的连接酶（E3） 并调节其转录活性。TP 53基因突变是EAC中最常见的事件。 LGD和HGD增加。有两种Grail同种型：RNF 128-Iso 1和RNF 128-Iso 2， 单独的发起人。Iso 1在BE/LGD中丰富，在HGD和EAC中显著减少，在 EAC细胞在用炎性（IFN-γ）或DNA损伤（H2 O2）剂处理后。此外，本发明还 我们发现，Iso 1的减少导致Iso 2糖基化形式的补偿性上调， 与WT TP 53相反，其稳定突变型TP 53 * 蛋白以增加克隆形成存活， 在永生化BE细胞中对抗炎药物（他汀类）的抗性。因此，了解 Grail同种型在BE/EAC中对野生型和TP 53 * 功能/稳定性的调节可能导致有效的 调节这一重要基因并预防Barrett's患者癌症发展的方法 化生

Abstract: Esophageal adenocarcinoma (EAC) is a deadly cancer that arises within a premalignant field called Barrett’s metaplasia (BE). EAC is increasing in incidence at a rate greater than any other cancer and is associated with gastroesophageal reflux disease (GERD) and obesity. Understanding which properties of Barrett’s metaplastic epithelium are associated with progression to EAC may potentially identify novel therapeutic strategies to prevent this cancer. We utilized the BE obtained from patients who progressed to high-grade dysplasia and EAC and performed RNAseq analysis for gene expression as well as isoform-specific mRNA analysis in non-dysplastic Barrett’s, low-grade dysplasia (LGD), high-grade dysplasia (HGD) to EAC. Importantly, we observe that transition from Barrett’s/LGD to HGD/EAC is associated with significant isoform changes within individual genes. This “isoform switching” is consistent with pathway analysis indicating splicing as the top pathway altered in BE progression to EAC. We observe that “isoform switching” coincides with the loss of protective mucins, increased inflammation, activation of ATM/DNA damage response pathway and increased H2AX staining in HGD and EAC. ATM activation was recently reported to regulate splicing. One of our top genes that demonstrate dramatic isoform switching is RNF128 or Grail, an ubiquitin ligase (E3) that functionally interacts with the N-terminus of wild type (WT) TP53 to target its degradation and modulate its transactivation activity. Mutations in the TP53 gene are the most frequent event in EAC and increase in LGD and HGD. There are two Grail isoforms: RNF128-Iso1 and RNF128-Iso2 that have separate promoters. Iso1 is abundant in BE/LGD, dramatically reduced in HGD and EAC and decreased in EAC cells following treatment with either inflammatory (-IFN) or DNA damaging (H2O2) agents. Additionally, we found that the reduction in Iso1 causes a compensatory upregulation of the glycosylated form of Iso2 protein, which, in contrast to WT TP53, stabilizes mutant TP53* protein to increase clonogenic survival and resistance to anti-inflammatory drugs (statins) in immortalized BE cells. Thus, understanding the role of Grail isoforms in the regulation of wild type and TP53* function/stability in BE/EAC may lead to effective methods to modulate this important gene and prevent cancer development in patients with Barrett’s metaplasia.