Kamps M P, Look A T, Baltimore D
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142.
Genes Dev. 1991 Mar;5(3):358-68. doi: 10.1101/gad.5.3.358.
The t(1;19) translocation that characterizes 25% of pediatric pre-B cell acute lymphoblastic leukemias (pre-B ALL) produces a chimeric gene, joining 5' sequences that encode a transcriptional activator domain of E2A with 3' sequences that, in part, encode a homeo box domain of a new gene called pbx1. Two E2A-pbx1 transcripts have been cloned. They encode the putative fusion proteins, p85(E2A-Pbx1) and p77(E2A-Pbx1), which differ in Pbx1 sequences alone, containing unique carboxyl termini whose sequences diverge after the Pbx1 homeo box. In this study, an antiserum to Pbx1 was used to investigate the identity and abundance of E2A-Pbx1 fusion proteins in both the pre-B ALL cell line, 697, and in cryopreserved leukemic bone marrow cells, obtained from six children with t(1;19)-positive pre-B ALL. Five species of E2A-Pbx1 proteins were identified in all cells containing t(1;19), two of which were indistinguishable from in vitro-translated p85(E2A-Pbx1) and p77(E2A-Pbx1). To assess the biological properties of p85(E2A-Pbx1) and p77(E2A-Pbx1) in fibroblasts, the cDNAs encoding these proteins were cloned into retroviral vectors, and each was introduced into NIH-3T3 cells. Both p85(E2A-Pbx1) and p77(E2A-Pbx1) are localized in the nucleus, and expression of either resulted in malignant conversion of NIH-3T3 cells as assayed by tumor formation in nude mice. When scored by focus formation, density-independent growth, and growth in agar assays, p77(E2A-Pbx1) was a much more potent transforming protein than was p85(E2A-Pbx1). Because subtle mutations in p85(E2A-Pbx1) converted its transforming activity into that of p77(E2A-Pbx1), we suggest that a sequence within the unique carboxyl terminus of p85(E2A-Pbx1) serves to negatively regulate its biochemical activity.
1;19易位存在于25%的儿童前B细胞急性淋巴细胞白血病（前B-ALL）中，它产生一种嵌合基因，将编码E2A转录激活结构域的5'序列与部分编码名为pbx1的新基因同源盒结构域的3'序列连接起来。已克隆出两种E2A-pbx1转录本。它们编码假定的融合蛋白p85（E2A-Pbx1）和p77（E2A-Pbx1），这两种蛋白仅在Pbx1序列上有所不同，包含独特的羧基末端，其序列在Pbx1同源盒之后出现分歧。在本研究中，使用针对Pbx1的抗血清来研究前B-ALL细胞系697以及从6名1;19阳性前B-ALL患儿获得的冻存白血病骨髓细胞中E2A-Pbx1融合蛋白的特性和丰度。在所有含有1;19易位的细胞中鉴定出了5种E2A-Pbx1蛋白，其中两种与体外翻译的p85（E2A-Pbx1）和p77（E2A-Pbx1）无法区分。为了评估p85（E2A-Pbx1）和p77（E2A-Pbx1）在成纤维细胞中的生物学特性，将编码这些蛋白的cDNA克隆到逆转录病毒载体中，并分别导入NIH-3T3细胞。p85（E2A-Pbx1）和p77（E2A-Pbx1）都定位于细胞核，并且通过裸鼠肿瘤形成实验检测发现，二者的表达均导致NIH-3T3细胞发生恶性转化。当通过集落形成、密度依赖性生长和琼脂培养实验进行评分时，p77（E2A-Pbx1）是一种比p85（E2A-Pbx1）更有效的转化蛋白。由于p85（E2A-Pbx1）中的微小突变将其转化活性转变为p77（E2A-Pbx1）的转化活性，我们认为p85（E2A-Pbx1）独特羧基末端内的一个序列起到负调控其生化活性的作用。
