DNA-binding specificity and trans-activating potential of the leukemia-associated E2A-hepatic leukemia factor fusion protein.

Abstract

Hybrid transcription factors, resulting from gene fusions in the wake of chromosomal translocations, have been implicated in leukemogenesis, but their precise contributions to oncogenic conversion remain unclear. The E2A-HLF fusion gene, formed by a t(17;19)(q22;p13) in childhood pro-B-cell acute lymphoid leukemia, encodes a hybrid protein that contains the trans-activation domain of E2A (E12/E47) linked to the bZIP DNA-binding and dimerization domain of hepatic leukemia factor (HLF). Here we report that both HLF and E2A-HLF bind to a 10-bp consensus sequence, 5'-GTTACGTAAT-3', with a core dyad-symmetric motif characteristic of the bZIP scissors-grip model of DNA binding. A probe containing this sequence bound chimeric E2A-HLF proteins in nuclear extracts of a leukemic cell line (UOC-B1) containing the t(17;19), as demonstrated by complexes supershifted with antibodies specific for amino-terminal epitopes of E2A or carboxyl-terminal eptiopes of HLF. E2A-HLF functioned as a potent trans activator of reporter gene expression from a plasmid that contained the consensus DNA-binding sequence. Interestingly, wild-type HLF was restricted in its capacity to act as a trans activator, functioning in human fetal kidney cells but not HepG2 hepatocarcinoma cells or NIH 3T3 mouse fibroblasts. The ability of the E2A-HLF hybrid protein to bind DNA in a sequence-specific manner and trans activate the expression of artificial reporter genes suggests that it could subvert transcriptional programs that normally control the growth, differentiation, and survival of lymphoid progenitor cells.