E2A-HLF-mediated cell transformation requires both the trans-activation domains of E2A and the leucine zipper dimerization domain of HLF.
The E2A-HLF fusion gene, formed by the t(17;19)(q22;p13) translocation in childhood acute pro-B-cell leukemia, encodes a hybrid protein that contains the paired trans-activation domains of E2A (E12/E47) linked to the basic region/leucine zipper DNA-binding and dimerization domain of hepatic leukemia factor (HLF). To assess the transforming potential of this novel gene, we introduced it into NIH 3T3 murine fibroblasts by using an expression vector that also contained the neomycin resistance gene. Cells selected for resistance to the neomycin analog G418 formed aberrant colonies in monolayer cultures, marked by increased cell density and altered morphology. Transfected cells also grew readily in soft agar, producing colonies whose sizes correlated with E2A-HLF expression levels. Subclones expanded from colonies with high levels of the protein reproducibly formed tumors in nude mice and grew to higher plateau-phase cell densities in reduced-serum conditions than did parental NIH 3T3 cells. By contrast, NIH 3T3 cells expressing mutant E2A-HLF proteins that lacked either of the bipartite E2A trans-activation domains or the HLF leucine zipper domain failed to show oncogenic properties, including anchorage-independent cell growth. Thus, both of the E2A trans-activation motifs and the HLF leucine zipper dimerization domain are essential for the transforming potential of the chimeric E2A-HLF protein, suggesting a model in which aberrant regulation of the expression pattern of downstream target genes contributes to leukemogenesis.
Yoshihara
T
T
Department of Experimental Oncology, St. Jude Children's Research Hospital, Memphis, Tennessee 38105, USA.
Inaba
T
T
Shapiro
L H
LH
Kato
J Y
JY
Look
A T
AT
D016475:3T3 Cells; D017502:Adenovirus E2 Proteins / Q000235:genetics* / Q000494:pharmacology; D000818:Animals; D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors; D002471:Cell Transformation, Neoplastic / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000494:pharmacology; D016350:Leucine Zippers / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D015514:Oncogene Proteins, Fusion / Q000235:genetics* / Q000494:pharmacology; D010957:Plasmids; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism
D017502:Adenovirus E2 Proteins; D050976:Basic-Leucine Zipper Transcription Factors; D004268:DNA-Binding Proteins; C489626:E2a-Hlf fusion protein, mouse; D015514:Oncogene Proteins, Fusion; D011993:Recombinant Fusion Proteins