Genomic structure and promoter activity of the E1AF gene, a member of the ETS oncogene family

doi:10.1016/j.bbrc.2005.11.024

Biochemical and Biophysical Research Communications

Volume 339, Issue 1, 6 January 2006, Pages 325-330

https://doi.org/10.1016/j.bbrc.2005.11.024 Get rights and content

Abstract

E1AF is a member of the ETS oncogene family and is thought to be a human homologue of mouse PEA3. We have isolated a genomic clone of E1AF and analyzed the promoter activity of its 5′-flanking region. We identified a variation in exon 1, which depends on the cell type. There was no typical TATA box in the 5′-flanking region, but putative binding sites of a number of transcription factors including PEA3 as well as CAAT boxes were seen. A luciferase reporter assay indicated that the 5′-flanking region possesses promoter activity. Northern blot studies demonstrated significant expression of the E1AF gene in restricted tissues such as the pituitary gland, placenta, and fetal kidney. Moreover, the E1AF promoter was activated by E1AF itself and estrogen receptor. These findings suggest that E1AF is a housekeeping gene, whose expression is controlled in specific tissues.

Section snippets

Materials and methods

Cells. Human prostatic adenocarcinoma PC3, human neuroblastoma GOTO, human oral squamous-cell carcinoma SAS, human embryo lung fibroblast MRC-5, and mouse mammary carcinoma FM3A (Health Science Research Resources Bank) were cultured at 37 °C with 5% CO₂ in Dulbecco’s modified Eagle’s medium (DMEM) with 5% fetal bovine serum and penicillin/streptomycin.

Isolation of genomic clones. A human genomic library of human peripheral blood cells (from Japanese Cancer Research Resources Bank, Tokyo, Japan)

Genome organization of the human E1AF gene

To obtain a genomic clone of E1AF, we screened a genomic library of human peripheral blood cells and a P1 phage system using primers corresponding to the 5′-end of E1AF cDNA [2] (see Materials and methods). The E1AF gene spanned approximately 30 kb and contained 14 exons as shown previously [17]. To clarify the start site, we carried out 5′-RACE (rapid amplification of the cDNA end) analysis using several cell lines such as PC3, SAS, MRC-5, and GOTO. Sequence analysis by comparing a nucleotide

Acknowledgments

We thank Dr. T. Matsuda for the expression constructs of estrogen receptor and Dr. Adachi for the genomic library. This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan.

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Cited by (6)

ETV1, 4 and 5: An oncogenic subfamily of ETS transcription factors
2012, Biochimica et Biophysica Acta - Reviews on Cancer
Citation Excerpt :
Together with the observed upregulation of PEA3 factors in breast cancer, these data implicate that PEA3 factor overexpression contributes to the causation of mammary tumors. Molecular studies suggest a vicious cycle: HER2/Neu activates the transcriptional activity of PEA3 factors [51,52], which in turn bind to the ETV4 and HER2/Neu gene promoters and stimulate their transcription [16,40,53]. This is controversial, since one report posits that ETV4 binds to and thereby represses the HER2/Neu gene promoter [54].
The homologous ETV1, ETV4 and ETV5 proteins form the PEA3 subfamily of ETS transcription factors. In Ewing tumors, chromosomal translocations affecting ETV1 or ETV4 are an underlying cause of carcinogenesis. Likewise, chromosomal rearrangements of the ETV1, ETV4 or ETV5 gene occur in prostate tumors and are thought to be one of the major driving forces in the genesis of prostate cancer. In addition, these three ETS proteins are implicated in melanomas, breast and other types of cancer. Complex posttranslational modifications govern the activity of PEA3 factors, which can promote cell proliferation, motility and invasion. Here, we review evidence for a role of ETV1, 4 and 5 as oncoproteins and describe modes of their action. Modulation of their activation or interaction with cofactors as well as inhibiting crucial target gene products may ultimately be exploited to treat various cancers that are dependent on the PEA3 group of ETS transcription factors.
Adenovirus E1A negatively regulates E1AF, an ets family of the protein
2007, Biochemical and Biophysical Research Communications
Citation Excerpt :
The N-terminal region and CR1 of E1A is known as the binding region for several cellular proteins such as CBP/p300, Myogenin and P/CAF [18], so it is prospected that these proteins are candidates for mediating factors of E1A repression. To elucidate the region in the E1AF promoter responsible for E1A, we carried out a luciferase assay using reporters including a series of deleted E1AF promoters [15]. The schematic structure of the E1AF promoter and its deleted derivatives used in this study are shown in Fig. 2A.
E1AF was first identified as a transcription factor that binds to enhancer motifs of the adenovirus E1A gene and is thought to be a human homologue of mouse PEA3, one of the ets oncoprotein families. Here we show the effect of E1A on the gene expression and function of E1AF. E1A repressed the activity of E1AF promoter, and the N-terminal region of E1A, which is involved in the oncogenic activity of E1A, was essential for this repression. The ability as a transcription factor of E1AF, as well as those of the other PEA3 subfamily members ER81 and ERM, was also repressed by E1A via the same oncogenic domain. Furthermore, E1AF repressed the transformation activity of E1A cooperating with E1B, whereas the other ets family Ets-1 enhanced this activity. These results suggest that E1AF is one of the targets of E1A.
Identification and characterization of novel ETV4 splice variants in prostate cancer
2023, Scientific Reports
Increased ETV4 expression correlates with estrogen-enhanced proliferation and invasiveness of cholangiocarcinoma cells
2018, Cancer Cell International
Tamoxifen downregulates Ets oncogene family members ETV4 and ETV5 in benign breast tissue: Implications for durable risk reduction
2011, Cancer Prevention Research
Identification of E1AF as a target gene of E2F1-induced apoptosis in response to DNA damage
2008, Journal of Biochemistry

¹: These two authors contributed equally to this work.

²: Present address: Division of Protein Information, Institute for Genome Research, The University of Tokushima, 3-18-15, Kuramoto-cho 770-8503, Tokushima-city, Japan.

View full text

Genomic structure and promoter activity of the E1AF gene, a member of the ETS oncogene family

Abstract

Section snippets

Materials and methods

Genome organization of the human E1AF gene

Acknowledgments

Biochem. Mol. Med.

Biochem. Biophys. Res. Commun.

J. Biol. Chem.

Gene

Genomics

J. Boil. Chem.

Cell

The ETS-domain: a new DNA-binding motif that recognizes a purine-rich core DNA sequence

Genes Dev.

Isolation of a cDNA encoding the adenovirus E1A enhancer binding protein: a new human member of the ets oncogene family

Nucleic Acids Res.

Molecular cloning and characterization of PEA3, a new member of the Ets oncogene family that is differentially expressed in mouse embryonic cells

Genes Dev.

A single ets-related transcription factor, E1AF, confers invasive phenotype on human cancer cells

Oncogene

Correlated expression of matrix metalloproteinases and ets family transcription factor E1A-F in invasive oral squamous-cell-carcinoma-derived cell lines

Am. J. Pathol.