Division of Hematology/Oncology, Cincinnati Children's Hospital Medical Center, University of Cincinnati, Faculty of Medicine, Cincinnati, OH, USA.
Genes Chromosomes Cancer. 2010 Mar;49(3):224-36. doi: 10.1002/gcc.20731.
The fusion oncoproteins PAX3-FOXO1 [t(2;13)(q35;q14)] and PAX7-FOXO1 [t(1;13)(p36;q14)] typify alveolar rhabdomyosarcoma (ARMS); however, 20-30% of cases lack these specific translocations. In this study, cytogenetic and/or molecular characterization to include FISH, reverse transcription polymerase chain reaction (RT-PCR), and sequencing analyses of five rhabdomyosarcomas [four ARMS and one embryonal rhabdomyosarcoma (ERMS)] with novel, recurrent t(2;2)(p23;q35) or t(2;8)(q35;q13) revealed that these noncanonical translocations fuse PAX3 to NCOA1 or NCOA2, respectively. The PAX3-NCOA1 and PAX3-NCOA2 transcripts encode chimeric proteins composed of the paired-box and homeodomain DNA-binding domains of PAX3, and the CID domain, the Q-rich region, and the activation domain 2 (AD2) domain of NCOA1 or NCOA2. To investigate the biological function of these recurrent variant translocations, the coding regions of PAX3-NCOA1 and PAX3-NCOA2 cDNA constructs were introduced into expression vectors with tetracycline-regulated expression. Both fusion proteins showed transforming activity in the soft-agar assay. Deletion of the AD2 portion of the PAX3-NCOA fusion proteins reduced the transforming activity of each chimeric protein. Similarly, but with greater impact, CID domain deletion fully abrogated the transforming activity of the chimeric protein. These studies (1) expand our knowledge of PAX3 variant translocations in RMS with identification of a novel PAX3-NCOA2 fusion, (2) show that both PAX3-NCOA1 and PAX3-NCOA2 represent recurrent RMS rearrangements, (3) confirm the transforming activity of both translocation events and demonstrate the essentiality of intact AD2 and CID domains for optimal transforming activity, and (4) provide alternative approaches (FISH and RT-PCR) for detecting PAX-NCOA fusions in nondividing cells of RMS. The latter could potentially be used as aids in diagnostically challenging cases.
PAX3-FOXO1 [t(2;13)(q35;q14)] 和 PAX7-FOXO1 [t(1;13)(p36;q14)] 融合癌蛋白是肺泡横纹肌肉瘤 (ARMS) 的典型特征；然而，20-30%的病例缺乏这些特定的易位。在这项研究中，通过 FISH、逆转录聚合酶链反应 (RT-PCR) 和对五个横纹肌肉瘤[四个 ARMS 和一个胚胎性横纹肌肉瘤 (ERMS)]的分子特征分析，包括五个横纹肌肉瘤[四个 ARMS 和一个胚胎性横纹肌肉瘤 (ERMS)]的核型分析和/或分子特征分析，揭示了这些非典型易位分别将 PAX3 融合到 NCOA1 或 NCOA2 上。PAX3-NCOA1 和 PAX3-NCOA2 转录本编码嵌合蛋白，由 PAX3 的配对盒和同源结构域 DNA 结合域、CID 结构域、富含 Q 的区域和 NCOA1 或 NCOA2 的激活结构域 2 (AD2) 结构域组成。为了研究这些反复出现的变体易位的生物学功能，将 PAX3-NCOA1 和 PAX3-NCOA2 cDNA 构建体的编码区引入具有四环素调控表达的表达载体中。两种融合蛋白在软琼脂测定中均具有转化活性。PAX3-NCOA 融合蛋白的 AD2 部分缺失降低了每种嵌合蛋白的转化活性。同样地，但影响更大，CID 结构域缺失完全消除了嵌合蛋白的转化活性。这些研究 (1) 扩展了我们对 RMS 中 PAX3 变体易位的认识，确定了一种新的 PAX3-NCOA2 融合，(2) 表明 PAX3-NCOA1 和 PAX3-NCOA2 均代表 RMS 的反复易位，(3) 证实了两种易位事件的转化活性，并证明了完整的 AD2 和 CID 结构域对于最佳转化活性的必要性，(4) 为 RMS 中非分裂细胞中 PAX-NCOA 融合的检测提供了替代方法（FISH 和 RT-PCR）。后者可能有助于诊断具有挑战性的病例。
