| Co-Investigator(Kenkyū-buntansha) |
KUROKAWA Mineo Tokyo University Hospital, research associate, 医学部・附属病院, 医員
OGAWA Seishi Tokyo University Hospital, assistant professor, 医学部・附属病院, 助手 (60292900)
TANAKA Tomoyuki Tokyo University Hospital, assistant professor, 医学部・附属病院, 助手 (50227154)
HIRAI Hisamaru Tokyo University Hospital, associate professor, 医学部・附属病院, 助教授 (90181130)
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| Research Abstract |
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
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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
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