Constitutive c-Myb amino-terminal phosphorylation and DNA binding activity uncoupled during entry and passage through the cell cycle.
The c-myb gene encodes a transcription factor that is central to hematopoietic cell growth. Phosphorylation of c-Myb by casein kinase 2 (CK2) at serines 11 and 12 has been variously implicated in the regulation of DNA binding. However, it is unclear when c-Myb phosphorylation at serines 11 and 12 occurs during the cell cycle and how this is regulated. We generated specific antisera that recognize phosphoserines 11 and 12 of c-Myb. C-Myb protein levels, extent of CK2 phosphorylation and DNA binding were then monitored following mitogenic stimulus and passage through the cell cycle in normal peripheral T-cells and the T leukemia cell line CCRF-CEM. We found that endogenous c-Myb is constitutively phosphorylated at serines 11 and 12. The amount of phosphorylated c-Myb correlates with DNA binding activity in cycling CEM cells but not upon entry of T-cells into the cell cycle. Exogenous expression of c-Myb with substitutions of serines 11 and 12 with glutamic acid or alanine had no effect on the transactivation of a c-Myb responsive reporter. These data strongly suggest that c-Myb is constitutively phosphorylated on serines 11 and 12 by CK2 or like activity and is not regulated during the cell cycle.
Cures
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Trescowthick Research Laboratories, Peter MacCallum Cancer Institute, Melbourne, Vic. Australia.
House
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Kanei-Ishii
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Kemp
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Ramsay
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D047390:Casein Kinase II; D002453:Cell Cycle; D004247:DNA / Q000378:metabolism*; D006801:Humans; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D020598:Proto-Oncogene Proteins c-myb / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
D020598:Proto-Oncogene Proteins c-myb; D012694:Serine; D004247:DNA; D047390:Casein Kinase II; D017346:Protein Serine-Threonine Kinases