LeBrun D P, Matthews B P, Feldman B J, Cleary M L
Department of Pathology, Queen's University, Kingston, Ontario, Canada.
Oncogene. 1997 Oct 23;15(17):2059-67. doi: 10.1038/sj.onc.1201367.
Oncogenic mutation of nuclear transcription factors often is associated with altered patterns of subcellular localization that may be of functional importance. The leukemogenic transcription factor gene E2A-PBX1 is created through fusion of the genes E2A and PBX1 as a result of t(1;19) in acute lymphoblastic leukemia. We evaluated subcellular localization patterns of E2A-PBX1 protein in transfected cells using immunofluorescence. Full-length E2A-PBX1 was exclusively nuclear and was concentrated in spherical domains denoted chimeric-E2A oncoprotein domains (CODs). In contrast, nuclear fluorescence for wild-type E2A or PBX1 proteins was diffuse. Enhanced concentrations of RNA polymerase II within many CODs and the requirement for an E2A-encoded activation domain suggested transcriptional relevance. However, in situ co-detection of nascent transcripts labeled with bromouridine failed to confirm altered transcriptional activity in relation to CODs. CODs also failed to co-localize with other proteins known to occupy functional nuclear compartments, including the transcription factor PML, the spliceosome-associated protein SC-35 and the adenovirus replication factor DBP, or with foci of DNA replication. Co-transfection of Hoxb7, a homeodomain protein capable of enhancing DNA binding by PBX1, impaired COD formation, suggesting that CODs contain E2A-PBX1 protein not associated with DNA. We conclude that, as a 'gain of function' phenomenon requiring protein elements from both E2A and PBX1, COD formation may be relevant to the biology of E2A-PBX1 in leukemogenesis.
核转录因子的致癌突变通常与亚细胞定位模式的改变有关，这可能具有功能重要性。白血病致癌转录因子基因E2A-PBX1是急性淋巴细胞白血病中t(1;19)导致E2A和PBX1基因融合而产生的。我们使用免疫荧光评估了转染细胞中E2A-PBX1蛋白的亚细胞定位模式。全长E2A-PBX1仅定位于细胞核，并集中在称为嵌合E2A癌蛋白结构域（COD）的球形结构域中。相比之下，野生型E2A或PBX1蛋白的核荧光是弥散的。许多COD中RNA聚合酶II浓度的增加以及对E2A编码的激活结构域的需求表明其与转录相关。然而，用溴尿苷标记的新生转录本的原位共检测未能证实与COD相关的转录活性改变。COD也未能与其他已知占据功能性核区室的蛋白质共定位，包括转录因子PML、剪接体相关蛋白SC-35和腺病毒复制因子DBP，也未与DNA复制位点共定位。能够增强PBX1与DNA结合的同源结构域蛋白Hoxb7的共转染会损害COD的形成，这表明COD包含与DNA不相关的E2A-PBX1蛋白。我们得出结论，作为一种需要E2A和PBX1两者蛋白质元件的“功能获得”现象，COD的形成可能与E2A-PBX1在白血病发生中的生物学特性相关。
