Department of Pediatrics, Memorial Sloan-Kettering Cancer Center, New York, USA.
Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, USA.
J Pathol. 2013 Apr;229(5):743-754. doi: 10.1002/path.4158. Epub 2013 Mar 5.
Oncogenic rearrangements of the TFE3 transcription factor gene are found in two distinct human cancers. These include ASPSCR1-TFE3 in all cases of alveolar soft part sarcoma (ASPS) and ASPSCR1-TFE3, PRCC-TFE3, SFPQ-TFE3 and others in a subset of paediatric and adult RCCs. Here we examined the functional properties of the ASPSCR1-TFE3 fusion oncoprotein, defined its target promoters on a genome-wide basis and performed a high-throughput RNA interference screen to identify which of its transcriptional targets contribute to cancer cell proliferation. We first confirmed that ASPSCR1-TFE3 has a predominantly nuclear localization and functions as a stronger transactivator than native TFE3. Genome-wide location analysis performed on the FU-UR-1 cell line, which expresses endogenous ASPSCR1-TFE3, identified 2193 genes bound by ASPSCR1-TFE3. Integration of these data with expression profiles of ASPS tumour samples and inducible cell lines expressing ASPSCR1-TFE3 defined a subset of 332 genes as putative up-regulated direct targets of ASPSCR1-TFE3, including MET (a previously known target gene) and 64 genes as down-regulated targets of ASPSCR1-TFE3. As validation of this approach to identify genuine ASPSCR1-TFE3 target genes, two up-regulated genes bound by ASPSCR1-TFE3, CYP17A1 and UPP1, were shown by multiple lines of evidence to be direct, endogenous targets of transactivation by ASPSCR1-TFE3. As the results indicated that ASPSCR1-TFE3 functions predominantly as a strong transcriptional activator, we hypothesized that a subset of its up-regulated direct targets mediate its oncogenic properties. We therefore chose 130 of these up-regulated direct target genes to study in high-throughput RNAi screens, using FU-UR-1 cells. In addition to MET, we provide evidence that 11 other ASPSCR1-TFE3 target genes contribute to the growth of ASPSCR1-TFE3-positive cells. Our data suggest new therapeutic possibilities for cancers driven by TFE3 fusions. More generally, this work establishes a combined integrated genomics/functional genomics strategy to dissect the biology of oncogenic, chimeric transcription factors.
TFE3 转录因子基因的致癌重排存在于两种不同的人类癌症中。这些癌症包括肺泡软组织肉瘤（ASPS）中所有的 ASPSCR1-TFE3，以及儿童和成人群体肾细胞癌（RCC）中的一部分 ASPSCR1-TFE3、PRCC-TFE3、SFPQ-TFE3 等。在这里，我们研究了 ASPSCR1-TFE3 融合致癌蛋白的功能特性，在全基因组范围内定义了其靶启动子，并进行了高通量 RNA 干扰筛选，以确定其哪些转录靶标有助于癌细胞增殖。我们首先证实 ASPSCR1-TFE3 主要定位于核内，并作为比天然 TFE3 更强的转录激活子发挥作用。在表达内源性 ASPSCR1-TFE3 的 FU-UR-1 细胞系上进行的全基因组定位分析，确定了 2193 个被 ASPSCR1-TFE3 结合的基因。将这些数据与 ASPS 肿瘤样本的表达谱和表达 ASPSCR1-TFE3 的诱导细胞系整合在一起，确定了 332 个基因作为 ASPSCR1-TFE3 的假定上调直接靶标，包括 MET（先前已知的靶基因）和 64 个作为 ASPSCR1-TFE3 下调靶标的基因。作为识别真正的 ASPSCR1-TFE3 靶基因的这种方法的验证，通过多条证据表明，两种被 ASPSCR1-TFE3 结合的上调基因 CYP17A1 和 UPP1 是 ASPSCR1-TFE3 转录激活的直接、内源性靶标。由于结果表明 ASPSCR1-TFE3 主要作为一种强转录激活因子发挥作用，我们假设其上调的直接靶标中的一部分介导其致癌特性。因此，我们选择了这些上调的直接靶标中的 130 个，在 FU-UR-1 细胞中进行高通量 RNAi 筛选研究。除了 MET，我们还提供了证据表明，ASPSCR1-TFE3 的另外 11 个靶基因有助于 ASPSCR1-TFE3 阳性细胞的生长。我们的数据为 TFE3 融合驱动的癌症提供了新的治疗可能性。更普遍地说，这项工作建立了一种综合的基因组学/功能基因组学策略，用于剖析致癌嵌合转录因子的生物学。
