| Budget Amount *help |
¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1993: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Research Abstract |
In order to identify a ganglioside-binding protein regulating the differentiation and the proliferation of human leukemia cells, we prepared a photoaffinity probe with a ganglioside derivative. When 12-O-tetradecamoylphorbol 13-acetate (TPA) was exogenously added to HL-60 cells, a protein with a molecular weight of 40,000 and an isoelectric point of 5.2-5.3, which was photolabeled with GM3 probe, was induced. This protein was induced when more than 5 X 10^<-9> M TPA was supplemented and expressed biphasically in the time course of TPA treatment. First, it was weakly expressed at 1-2 h after the treatment. Second, it was strongly exoressed at 24 h after the treatment and the expression was maintained for four days as long as we examined. The 40 K protein was a soluble protein. The induction of the 40 K protein was also observed in a human acute monocytic leukemia cell line, THP-1. When TPA was added to THP-1 cells, only adherent cells expressed the protein. This result suggests that the GM3 binding protein is involved in the adherence to substance among differentiation phenotypes induced by TPA.Furthermore, we purified the 40 K protein from THP-1 cells using GM3 affinity chromatography.
Photoaffinity labeling of rat liver GM1 synthase, a beta-galactosyltransferase, revealed a protein with a molecular weight of 40,000 and an isoelectric point of 4.5 which comigrated with the enzyme activity.
We also succeeded in labeling Man6P/IGF-II receptor with a phosphomannan probe. The observation IGF-II didnot inhibit the affinity labeling confirmed that the site for Man6P was different from that for IGF-II.
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