Exogenous cdk4 overcomes reduced cdk4 RNA and inhibition of G1 progression in hematopoietic cells expressing a dominant-negative CBF - a model for overcoming inhibition of proliferation by CBF oncoproteins

Core Binding Factor (CBF) is required for the development of definitive hematopoiesis, and the CBF oncoproteins AML1-ETO, TEL-AML1, and CBF beta-SMMHC are commonly expressed in subsets of acute leukemia. CBF beta-SMMHC slows the G1 to S cell cycle transition in hematopoietic cells, but the mechanism of this effect is uncertain, We have sought to determine whether inhibition of CBF-mediated trans-activation is sufficient to stow proliferation. We demonstrate that activation of KRAB-AML1-ER, a protein containing the AML1 DNA-binding domain, the KRAB repression domain, and the Estrogen receptor ligand binding domain, also slows G1, if its DNA-binding domain is intact, Also, exogenous AML1 overcame CBF beta-SMMHC-induced inhibition of proliferation, Representational difference analysis (RDA) identified cdk4 RNA expression as an early target of KRAB-AML1 activation. Inhibition of CBF activities by KRAB-AML1-ER or CBF beta-SMMHC rapidly reduced endogenous cdk4 mRNA levels, even in cells proliferating at or near control rates as a result of exogenous cdk4 expression, Over-expression of cdk4, especially a variant which cannot bind p16(INK4a) overcame cell cycle inhibition resulting from activation of KRAB-AML1-ER, although cdk4 did not accelerate proliferation when expressed alone. These findings indicate that mutations which alter the expression of G1 regulatory proteins can overcome inhibition of proliferation by CBF oncoproteins.