METHODS: We established three transfectants of cell lines expressing different types of Z-fusion proteins, and analyzed their gene expression profile (GEP) by RNA-seq. We also analyzed the GEP of clinical ALL samples using our previous RNA-seq data of 323 Japanese ALL patients. We selected upregulated genes in both Z-fusion gene-expressing transfectants and Z-fusion gene-positive ALL samples, and investigated the binding of Z-fusion proteins to regulatory regions of the candidate genes by ChIP-qPCR.
RESULTS: We selected six commonly upregulated genes. After the investigation by ChIP-qPCR, we finally identified CREB5 and RGS1 as direct and common target genes. RGS1 is an inhibitor of CXCL12-CXCR4 signaling that is required for the homing of hematopoietic progenitor cells to the bone marrow microenvironment and development of B cells. Consistent with this, Z-fusion gene transfectants showed impaired migration toward CXCL12.
CONCLUSIONS: We identified CREB5 and RGS1 as direct and common transcriptional targets of Z-fusion proteins. The present results provide novel insight into the aberrant transcriptional regulation by Z-fusion proteins.
方法:我们建立了三种表达不同类型Z-融合蛋白的细胞系的转染子,并通过RNA-seq分析其基因表达谱(GEP)。我们还使用我们先前的323名日本ALL患者的RNA-seq数据分析了临床ALL样品的GEP。我们选择了表达Z融合基因的转染子和Z融合基因阳性ALL样品中的上调基因,并通过ChIP-qPCR研究了Z融合蛋白与候选基因调控区的结合。
结果:我们选择了6个常见的上调基因。经过ChIP-qPCR的调查,我们最终确定CREB5和RGS1为直接和共同的靶基因。RGS1是CXCL12-CXCR4信号传导的抑制剂,是造血祖细胞归巢到骨髓微环境和B细胞发育所必需的。与此一致,Z-融合基因转染子显示向CXCL12的迁移受损。
结论:我们确定CREB5和RGS1是Z-融合蛋白的直接和共同的转录靶标。本结果为Z-融合蛋白的异常转录调控提供了新的见解。