The molecular function of the oncogenic NAB2-STAT6 fusion protein

致癌NAB2-STAT6融合蛋白的分子功能

• 批准号: 10314387
• 负责人: Connor Mackenzie Hill
• 金额: $ 4.6万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-12-01 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10314387
• 关键词: molecular; function; oncogenic; NAB2; STAT6

Project Summary Solitary Fibrous Tumors (SFTs) are a mesenchymal tumor type that affects an estimated 10,000 Americans each year. 10-20% of these tumors become malignant and unresponsive to treatments. The pathogenesis of SFTs is currently unknown, as there are no recurring mutations in known tumor suppressors genes or oncogenes. The only recurring mutation identified in SFTs is a gene fusion between NAB2-STAT6 that results in a fusion protein. NAB2 and STAT6 are both transcription regulators. NAB2 is repressor of early growth response transcription factors (EGR1/2) and STAT6 is an activator of transcriptional programs in response to cytokines. Both proteins also contribute to enhancer activation. Despite both proteins' known functions, how NAB2-STAT6 affects gene expression in SFTs is unknown. Using the gene set enrichment analysis (GSEA) of a large microarray data set of SFTs we have found that the expression of both NAB2 and EGR1 targets were significantly upregulated in SFTs. However, the expression of STAT6 targets was unchanged. We expressed NAB2-STAT6 and performed ChIP-seq analysis and found that NAB2-STAT6 localizes to distal active transcriptional enhancers. Lastly, we analyzed RNA-seq and saw that almost 2000 genes were differentially expressed in Malignant tumors vs Benign tumors including several genes, which are regulated by well characterized transcriptional enhancers. We hypothesize that NAB2- STAT6 aberrantly activates EGR1 targets to increase proliferation and highjacks the activity of transcriptional enhancers to promote malignancy. We have generated the intra-chromosomal inversion responsible for NAB2-STAT6 expression in the benign lung fibroblast IMR90 cell line, which replicates the mesenchymal origin of SFTs using CRISPR-Cas9. In Aim 1 we will establish NAB2-STAT6’s role in directing aberrant gene expression. We will characterize the genomic binding profile of NAB2-STAT6 as well as its effect on gene expression and proliferation. In Aim 2 we will investigate the ability of NAB2-STAT6 to reprogram transcriptional enhancers to promote malignancy. First, we will develop an inducible NAB2-STAT6 system to measure the ability of NAB2-STAT6 to reprogram transcriptional enhancers. Then, we will validate our results in primary SFTs and examine differences in malignant vs benign tumors. Finally, we will examine the ability of transcriptional enhancer changes to affect gene expression through analysis of RNA-seq of primary SFTs. Overall, these aims will establish the function of NAB2-STAT6 in promoting tumorigenesis in SFTs

项目摘要 孤立性纤维性肿瘤(SFT)是一种间叶性肿瘤，估计影响10,000人。 每年都有美国人。这些肿瘤中有10%-20%变得恶性，对治疗没有反应。这个 SFTS的发病机制目前尚不清楚，因为在已知的肿瘤抑制基因中没有复发的突变 基因或致癌基因。在SFTS中发现的唯一反复发生的突变是NAB2-STAT6之间的基因融合 形成一种融合蛋白。NAB2和STAT6都是转录调控因子。NAB2是早期生长的抑制因子 反应转录因子(Egr1/2)和STAT6是转录程序的激活物 细胞因子。这两种蛋白质也有助于增强启动子的激活。尽管这两种蛋白质的功能都已知，但如何 NAB2-STAT6对SFT基因表达的影响尚不清楚。 利用对SFT的大型微阵列数据集的基因集浓缩分析(GSEA)，我们发现 在SFT中，NAB2和Egr1靶基因的表达均显著上调。然而， STAT6靶基因的表达没有变化。我们表达了NAB2-STAT6并进行了CHIP-SEQ分析 发现NAB2-STAT6定位于远端的活性转录增强子。最后，我们对rna-seq进行了分析。 发现近2000个基因在恶性肿瘤和良性肿瘤中有差异表达，包括 几个基因，它们由特征明确的转录增强子调控。我们假设NAB2- STAT6异常激活Egr1靶点促进增殖并上调转录活性 促进恶性肿瘤的增强剂。 我们已经在染色体内产生了导致NAB2-STAT6表达的倒位 良性肺成纤维细胞系IMR90，使用CRISPR-Cas9复制SFT的间充质来源。在……里面 目的1确定NAB2-STAT6‘S在指导基因异常表达中的作用。我们将描述 NAB2-STAT6的基因组结合谱及其对基因表达和增殖的影响。在《目标2》中，我们 将研究NAB2-STAT6重新编程转录增强子以促进恶性肿瘤的能力。第一, 我们将开发一个可诱导的NAB2-STAT6系统来测量NAB2-STAT6的重编程能力 转录增强剂。然后，我们将验证我们在主要SFT中的结果，并检查 恶性肿瘤与良性肿瘤的对比。最后，我们将研究转录增强子的变化对 通过对原代SFT的RNA-seq分析进行基因表达。总体而言，这些目标将确立 NAB2-STAT6在促进SFT肿瘤发生中的作用