Expression of an activated raf transgene accelerated the terminal myeloid differentiation of HL-60 human promyelocytic leukemia cells induced by retinoic acid. A similar result was obtained when 1,25-dihydroxy vitamin D3 was used to induce monocytic differentiation. The stable transfectants were derived by transfecting HL-60 cells with DNA encoding an N-terminal truncated raf-1 protein. In normal HL-60 cells retinoic acid is known to induce a CSF-1 dependent metabolic cascade culminating in GO arrest and phenotypic conversion. Early in this cascade, expression of the RB tumor suppressor gene product is down regulated. A progressive redistribution of the form of the protein from largely hyper-phosphorylated protein to the hypo-phosphorylated form begins later with GO arrest and differentiation. In the activated raf transfected cells, RB down regulation occurred more rapidly, consistent with accelerated differentiation. But the conversion to the hypo-phosphorylated form was not accelerated and occurred after GO arrest and phenotypic conversion to myeloid differentiated cells. Thus raf activation appears to be a component of the retinoic acid induced metabolic cascade culminating in terminal differentiation. In this cascade raf activation promotes RB down regulation. The data are consistent with a model in which raf is an effector of this CSF-1 dependent metabolic cascade which culminates in terminal cell differentiation, and RB down regulation is one of the downstream consequences of raf action. Furthermore they indicate that RB down regulation may be an essential component of the cellular processes causing GO arrest and differentiation, but RB hypo-phosphorylation is more likely a consequence thereof and not a cause.
KeywordsRetinoic Acid Aberrant Crypt Focus Normal Human Keratinocytes Retinyl Palmitate Perillyl Alcohol
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