EVI-1 zinc finger protein works as a transcriptional activator via binding to a consensus sequence of GACAAGATAAGATAAN1-28 CTCATCTTC.
Previously, the DNA-binding consensus sequences for domains 1 (GACAAGATAAGATAA) and 2 (GAAGATGAG) of the EVI-1 protein were identified using GST fusion proteins of each domain in binding and amplification reactions. We have utilized full-length EVI-1 protein to confirm these consensus sequences and determine the spacial and orientation requirements for binding. Our data demonstrate that full-length EVI-1 can independently bind the consensus sequences in gel mobility shift assays. In binding and amplification reactions only the domain 1 consensus sequence (D1-CONS) was obtained with full-length EVI-1 protein. However, by using constructs in which D1-CONS was anchored, products were obtained in which the domain 2 consensus sequence (D2-CONS) was observed with the spacing and orientation of GACAAGATAATATAAN1-28 CTCATCTTC. Using this consensus sequence we show that EVI-1 can activate transcription from reporter constructs. No transcriptional activation was seen with the reporter construct containing D1-CONS alone while activation was seen with the construct-containing D2-CONS alone. These results indicate that the EVI-1 protein works as a transcriptional activator and the binding of the domain 2 with D2-CONS is essential for its activation.
Morishita
K
K
Biology Division, National Cancer Center Research Institute, Tokyo, Japan.
Suzukawa
K
K
Taki
T
T
Ihle
J N
JN
Yokota
J
J
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D016384:Consensus Sequence / Q000502:physiology*; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D005260:Female; D006801:Humans; D000074008:MDS1 and EVI1 Complex Locus Protein; D051379:Mice; D008969:Molecular Sequence Data; D011519:Proto-Oncogenes; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D016335:Zinc Fingers / Q000502:physiology*
D004268:DNA-Binding Proteins; D000074008:MDS1 and EVI1 Complex Locus Protein; C063575:MECOM protein, human; C499305:Mecom protein, mouse; D014157:Transcription Factors; D004247:DNA