The Evi-1 proto-oncogene encodes a transcriptional repressor activity associated with transformation

Abstract

The myeloid transforming gene Evi-1 encodes a protein with two zinc ®nger domains, designated ZF1 and ZF2, with distinct DNA binding speci®cities. For the ®rst time we demonstrate that Evi-1 has transcriptional repressor activity which is directly proportional to the amount of Evi-1 protein in cells. Repression has been observed with two distinct promoters: the minimal HSV-1 tk promoter and a VP16 inducible adenovirus E1b minimal promoter. Optimal repression is DNA binding dependent and is mediated by either ZF1 or a heterologous GAL4 DNA binding domain (GAL4DBD) but is signi®cantly less e cient through the ZF2 binding site. Both GAL4DBD/ Evi-1 fusion and non-fusion proteins have been used to map the repressor activity to a proline-rich region located within amino acids 514 ± 724 between the ZF1 and ZF2 domains. Constitutive expression of mutant proteins lacking the repressor domain are defective for transformation of Rat1 ®broblasts demonstrating that this region is required for the oncogenic activity of the Evi-1 protein. These studies show that the Evi-1 gene encodes a transcriptional repressor and has important implications for the mechanism of action of the Evi-1 protein both in development and in the progression of some myeloid leukaemias.

The EVI1 gene in myeloid leukemia

“…32 In contrast, studies using CAT reporters that contained either the first or second EVI1 target sites upstream of the HSVtk promoter found that EVI1 dramatically expressed the activity of these reporters. 28,33 More recently, studies by Bartholomew et al 34 showed that EVI1 represses synthetic promoters in which the zinc fingers consensus site was cloned upstream of the herpes simplex virus (HSV) thymidine kinase (tk) promoter or of the adenovirus minimal promoter, confirming the role of EVI1 as transcriptional repressor. In addition, these investigators showed that the repression function is localized in a stretch of proline-rich residues located between the two groups of zinc fingers (Figure 1a), and it can be completely abolished by an internal deletion spanning the proline-rich stretch.…”

“…In addition, these investigators showed that the repression function is localized in a stretch of proline-rich residues located between the two groups of zinc fingers (Figure 1a), and it can be completely abolished by an internal deletion spanning the proline-rich stretch. 34 …”

“…This protein is generally considered a repressor, and a stretch of amino acid residues necessary for repression has been identified. 34 We have recently compared the transcriptional effects of EVI1 and MDS1/EVI1 using a genomic promoter containing several repeats of the AGATA sequence. By using reporter gene assays, we determined that whereas MDS1/EVI1 is an activator of the promoter, EVI1 is a strong repressor.…”

Tumor-specific adenoviral gene therapy: transcriptional repression of gene expression by utilizing p53-signal transduction pathways

“…[18][19][20][21][22][23][24][25][26]55,56 The KRAB-domain represses the activity of CMV-promoter through direct communications with TATA box-dependent basal transcription machinery, 18,55 whereas mSIN3B interacts with essential DNAbinding proteins that place it in a promoter context in which it can repress transcription by recruiting proteins that condense chromatin by histone deacetylation. 19 In contrast to KRAB-A or mSIN3B, the Zik1-repressor domain does not repress gene transcription from the CMV gal -promoter and some of the other repressor domains showed only weak repression of CMV galactivity.…”

“…Expression vectors for repressor fusion proteins of GAL4-DBD were kindly provided by: L Lania, Neaples, Italy (KRAB-A), 18 RA DePinho Boston, USA (m-SIN3B-SF and m-SIN3B-LF), 19 DJ Hall, Philadelphia, PA, USA (ZF87/MAZ), 20 K Bomsztyk, Seattle, USA (ZIK1), 21 LM Staudt, Bethesda, MD, USA (BCL-6), 22 C Bartholomew, Glasgow, Scotland (EVI-1), 23 Y Shi, Boston, USA (YY), 24 H-S Ro, Halifax Canada (AEBP2) 25 and A Bird, Edinburgh, UK (MeCP2 170-310 and MeCP2 209-492). 26 …”