Yamamoto K, Seto M, Akao Y, Iida S, Nakazawa S, Oshimura M, Takahashi T, Ueda R
Laboratory of Chemotherapy, Aichi Cancer Center Research Institute, Nagoya, Japan.
Oncogene. 1993 Feb;8(2):479-85.
We have previously demonstrated that the breakpoints of t(11;19)(q23;p13) leukemias are within 360 kb of the CD3 gene. One of the phage clones, 6n, which was isolated from the yeast artificial chromosome clone yB22B2 containing CD3, was found to be within 60 kb of t(11;19) breakpoints. In this study, gene walking was conducted and two phage clones (lambda Hp8-3 and lambda Hp23-13) were isolated from a human placenta genomic library. Southern blot analysis with a genomic probe from lambda Hp8-3 detected gene rearrangements in t(4;11) and t(11;19) cell lines with BamHI digestion. Subsequently, using reiterated sequence-free probes from both ends of 6n that detected transcriptional units in various hematopoietic cells, we isolated cDNA clones. These cDNA clones were classified into two groups (designated MLL-a and MLL-b), which do not hybridize to each other. Northern blot analysis with MLL-a cDNA detected 15-, 14- and 12-kb mRNAs, while MLL-b detected the additional 9.7- and 5-kb mRNAs in peripheral blood lymphocytes. MLL-b cDNA detected a truncated form of 12.5-kb mRNA in t(4;11) cell lines and a truncated form of 10-kb or 9.2-kb mRNA in t(11;19) cell lines. MLL-a did not demonstrate a truncated form of mRNA, but the stronger 14-kb signal was noted in t(4;11) cells, while this signal was very weak in t(11;19) cells. By Southern blot analysis, MLL-b cDNA detected gene rearrangement in cell lines with t(4;11) and t(11;19), whereas MLL-a did not. Furthermore, chimeric cDNA clones were isolated from cDNA libraries of t(4;11) and t(11;19) cell lines with a MLL-b cDNA probe. These results indicate that the MLL-b cDNA is derived from the common target gene involved in 11q23 translocation with 4q21 or 19p13.
我们之前已经证明，t(11;19)(q23;p13)白血病的断点位于CD3基因的360 kb范围内。从包含CD3的酵母人工染色体克隆yB22B2中分离出的一个噬菌体克隆6n，被发现位于t(11;19)断点的60 kb范围内。在本研究中，进行了基因步移，并从人胎盘基因组文库中分离出两个噬菌体克隆（λHp8 - 3和λHp23 - 13）。用来自λHp8 - 3的基因组探针进行Southern印迹分析，用BamHI消化检测t(4;11)和t(11;19)细胞系中的基因重排。随后，使用来自6n两端的无重复序列探针，这些探针在各种造血细胞中检测转录单位，我们分离出了cDNA克隆。这些cDNA克隆被分为两组（命名为MLL - a和MLL - b），它们彼此不杂交。用MLL - a cDNA进行Northern印迹分析检测到15 kb、14 kb和12 kb的mRNA，而MLL - b在外周血淋巴细胞中检测到另外的9.7 kb和5 kb的mRNA。MLL - b cDNA在t(4;11)细胞系中检测到12.5 kb mRNA的截短形式，在t(11;19)细胞系中检测到10 kb或9.2 kb mRNA的截短形式。MLL - a未显示mRNA的截短形式，但在t(4;11)细胞中观察到较强的14 kb信号，而在t(11;19)细胞中该信号非常弱。通过Southern印迹分析，MLL - b cDNA在t(4;11)和t(11;19)细胞系中检测到基因重排，而MLL - a未检测到。此外，用MLL - b cDNA探针从t(4;11)和t(11;19)细胞系的cDNA文库中分离出嵌合cDNA克隆。这些结果表明，MLL - b cDNA源自与4q21或19p13发生11q23易位相关的共同靶基因。
