In this study, the abilities of constitutive and conditional forms of the three Raf kinases to abrogate the cytokine dependency of FDC-P1 cells were examined. The constitutively active forms (⌬) of all three Raf kinases were fused to the hormone-binding domain of the estrogen receptor (ER), rendering their activities conditionally dependent upon exogenous ␤-estradiol. The vast majority of ⌬Raf:ER-infected FDC-P1 cells remained cytokinedependent; however, cells were obtained at low frequency in which expression of ⌬Raf:ER abrogated cytokine dependency. Isoform specific differences between the Raf kinases were observed as cytokine-independent cells were obtained more frequently from ⌬A-Raf:ER than either ⌬Raf-1:ER or ⌬B-Raf:ER infected cells. To determine whether the regulatory phosphorylation sites in the Raf proteins were necessary for abrogation of cytokine dependency, they were changed by site-directed mutagenesis. Substitution with phenylalanine eliminated the transforming ability of the ⌬B-Raf:ER and ⌬Raf-1:ER kinases.

No abstract

No abstract

No abstract

No abstract