Analysis of transcription factor abnormality in the progression of preleukemic state.

白血病前期状态进展中转录因子异常的分析。

• 批准号: 08671215
• 负责人: MITANI Kinuko
• 金额: $ 1.6万
• 依托单位: Tokyo University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08671215/
• 关键词: RNA polymerase II elongation factor; elongation factor; MEN; t(11 ; 19); leukemia; AP-1 activity; 骨髄異形成症候群; t(11;19)(q.23;P13.1); MLL; 転写因子; 伸長因子

We have cloned an MIL/MEN(also termed as ELL)chimeric gene generated by the t(11 ; 19) (q23 ; p13.1), which is usually found in acute myeloid leukemia or myelodysplastic syndrome-derived leukemia. The MIL/MEN fusion protein contains the amino-terminal half of MLL including AT hook motifs which is fused to the MEN protein with a lysine-rich sequence(K.Mitani, et al. BLOOD 85 ; 2017,1995). Thus, the Mll/MEN fusion protein could be a chimeric transcription factor. Polyclonal antibodies to MLL and MEN were raised in rabbits against GST fusion of MLL(645-974 a.a.)and maltose-binding fusion of MEN(100-179 a.a.), respectively. These antibodies detected the MLL/MEN fusion protein of 230 kD,truncated form of the MLL Without the zinc finger domain and its downstream sequence(tMLL)of 180kD,and MEN protein of 70kD in Cos1 cells transfected with the corresponding cDNAs. Using immunostaining assay and subcellular fractionation of Cos1 cells expressing the MLL/MEN fusion, tMLL,and MEN proteins, we ha … More ve demonstrated that all these proteins exist in the nucleus of the cells.Conaway et al.have reported that the MEN is an RNA polymerase II elongation factor (SCIENCE 271 ; 1873,1996). The function of another elongation factor, elongin, is known to be inhibited by VHL tumor suppressor protein in vitro, suggesting the possible relationship of aberrant transcriptional elongation to oncogenesis. To demonstrate the transforming activity of the MEN protein, the MEN cDNA was introduced retrovirally into Ratl cells. Men-overexpressing cells acquired capacity of anchorage independent growth. In addition, the growth factor requirement was decreased in these cells. However, cells expressing a deletion mutant of MEN lacking the lysine-rich region did not exhibit such biological abilities. The c-Fos protein expression and AP-1 activity were elevated in the MEN-expressing cells, which might be part of the responsible mechanism for the transformation. The c-fos mRNA,the expression of which is known to be regulated partly at the stage of transcriptional elongation, appeared earlier in the MEN-expressing cells than in cells transfected with an empty vector or the deletion mutant lacking the lysine-rich region after the stimulation with epidermal growth factor. Overexpression or aberrant expression of MEN may play an important role in the leukemic tranfformation of stem cell disease. Less

我们克隆了一个由t(11；19)(q23；p13.1)产生的MIL/MEN(也称为ELL)嵌合基因，这种嵌合基因通常见于急性髓系白血病或骨髓增生异常综合征衍生的白血病。MIL/MEN融合蛋白含有MLL的氨基末端半部分，包括AT钩状基序，该基序与MEN蛋白融合，具有丰富的赖氨酸序列(K.Mitani等人)。血液85；2017、1995)。因此，MLL/MEN融合蛋白可能是一种嵌合转录因子。分别制备了抗MLL和MEN的多克隆抗体，分别对抗MLL的GST融合(645-974 A.A.)和MEN的麦芽糖结合融合(100-179 A.A.)。这些抗体在COS1细胞中检测到230kD的MLL/MEN融合蛋白、缺失锌指结构域的截短型MLL及其下游180kD的序列(TMLL)和70kD的MEN蛋白。通过免疫染色和亚细胞分离表达mll/man融合蛋白、tmll和man蛋白，我们得到了…。更多的证据表明，所有这些蛋白质都存在于细胞核中。康纳韦等人报告说，这些人是一种RNA聚合酶II延长因子(Science 271；1873,1996)。另一种延伸素的功能已知在体外被VHL肿瘤抑制蛋白抑制，提示异常转录延长素可能与肿瘤的发生有关。为了证明MENS蛋白的转化活性，将MENS基因逆转录病毒导入Ratl细胞。男性过表达细胞获得了锚定独立生长的能力。此外，这些细胞对生长因子的需求也减少了。然而，缺乏赖氨酸富集区的男性表达缺失突变的细胞并没有表现出这样的生物学能力。在表达MAN的细胞中，c-Fos蛋白表达和AP-1活性升高，这可能是导致转化的部分机制。C-fos m RNA的表达在转录延伸阶段被部分调控，在表皮生长因子刺激后，c-fos m RNA在表达MAN的细胞中比空载体或缺失赖氨酸富集区的细胞更早出现。男性的过度表达或异常表达可能在干细胞疾病的白血病转化中起重要作用。较少

项目成果

Mitani, K., Hangaishi, A., Imamura, N., Miyagawa, K., Ogawa, S., Kanda, Y., Yazaki, Y., Hirai, H.: "No concomitant occurrence of the N-ras and p53 gene mutations in myelodysplastic syndrome." Leukemia. 11. 863-865 (1997)

Mitani, K.、Hangaishi, A.、Imamura, N.、Miyakawa, K.、Okawa, S.、Kanda, Y.、Yazaki, Y.、Hirai, H.：“N-ras 和

Hirai, H., Ogawa, S., Hangaishi, A., Takahashi, T., Hurokawa, M., Mitani, K., Ueda, R., Yazaki, Y.: "Recent progress in molecular mechanisms of leukemogenesis : The cyclin-dependent kinase 4-inhibitor gene in human leukemias." Leukemia. 11. 358-360 (1997)

Hirai, H.、Okawa, S.、Hangaishi, A.、Takahashi, T.、Hurokawa, M.、Mitani, K.、Ueda, R.、Yazaki, Y.：“白血病发生分子机制的最新进展：

Tanaka, K,Tanaka, T,Kurokawa, M,Imai, Y,Ogawa, S,Mitani, K,Yazaki, Y,and Hirai, H.: "The AML1/ETO(MTG8)and AML1/Evi-1 leukemia-associated chimeric oncoproteins accumulate PEBP2beta(CBFbeta)in the nucleus more efficiently than wild-type AML1." Blood. (in p

田中 K、田中 T、黑川 M、今井 Y、小川 S、三谷 K、矢崎 Y 和平井 H.：“AML1/ETO(MTG8) 和 AML1/Evi-1 白血病 -

Sasaki K: "TPO/c-MPL ligand induces tyrosine phosphorylation multiple cellular proteins including proto-oncogene produckts,VAV and c-CBL,and RAS signaling molecules." Biochem.Biophys.Res.Commun.216. 338-347 (1996)

Sasaki K：“TPO/c-MPL 配体诱导多种细胞蛋白酪氨酸磷酸化，包括原癌基因产物、VAV 和 c-CBL 以及 RAS 信号分子。”

Yamagata T: "Triple synergism of human T-lymphotropic virus type l-encoded Tax,GATA-binding protein,and AP-1 is required for constitutive expression of the interleukin-5 gene in adult T-cell leukemia cells." Mol.Cell.Biol.17. 4272-4281 (1997)

Yamagata T：“成人 T 细胞白血病细胞中白细胞介素 5 基因的组成型表达需要人类 T 淋巴细胞病毒 L 型编码的 Tax、GATA 结合蛋白和 AP-1 的三重协同作用。”