Abstract
BackgroundETV4 is one of the ETS proteins overexpressed in prostate cancer (PC) as a result of recurrent chromosomal translocations. In human prostate cell lines, ETV4 promotes migration, invasion, and proliferation; however, its role in PC has been unclear. In this study, we have explored the effects of ETV4 expression in the prostate in a novel transgenic mouse model.MethodsWe have created a mouse model with prostate-specific expression of ETV4 (ETV4 mice). By histochemical and molecular analysis, we have investigated in these engineered mice the expression of p21, p27, and p53. The implications of our in vivo findings have been further investigated in human cells lines by chromatin-immunoprecipitation (ChIP) and luciferase assays.ResultsETV4 mice, from two independent transgenic lines, have increased cell proliferation in their prostate and two-thirds of them, by the age of 10 months, developed mouse prostatic intraepithelial neoplasia (mPIN). In these mice, cdkn1a and its p21 protein product were reduced compared to controls; p27 protein was also reduced. By ChIP assay in human prostate cell lines, we show that ETV4 binds to a specific site (-704/-696 bp upstream of the transcription start) in the CDKN1A promoter that was proven, by luciferase assay, to be functionally competent. ETV4 further controls CDKN1A expression by downregulating p53 protein: this reduction of p53 was confirmed in vivo in ETV4 mice.ConclusionsETV4 overexpression results in the development of mPIN but not in progression to cancer. ETV4 increases prostate cell proliferation through multiple mechanisms, including downregulation of CDKN1A and its p21 protein product: this in turn is mediated through direct binding of ETV4 to the CDKN1A promoter and through the ETV4-mediated decrease of p53. This multi-faceted role of ETV4 in prostate cancer makes it a potential target for novel therapeutic approaches that could be explored in this ETV4 transgenic model.
Highlights
ETS variant gene 4 (ETV4) is one of the ETS proteins overexpressed in prostate cancer (PC) as a result of recurrent chromosomal translocations
To assess in vivo the pathogenic role of ETV4 in prostate, we engineered a vector that expresses this ETV4-encoded shortened protein [34] under the control of a modified rat probasin promoter (PB) (Fig. 1a, b) that is able to drive an androgen-inducible prostate-specific expression [56]. By using this Rat probasin promoter (PB)-ETV4 vector, we have obtained six founders in the FVB strain: three of these were able to transmit the transgene to their progeny and two of them expressed the exogenous human ETV4 (hETV4) in the prostate of the derived mouse lines (ETV4-A, ETV4-B) (Fig. 1c, d)
ETV4 affects cell proliferation by modulating of cyclin-dependent kinase inhibitor 1A (Cdkn1a) and cyclin-dependent kinase inhibitor 1B (Cdkn1b) in vivo and in vitro The role of ETV4 in proliferation has not been broadly studied; we have previously demonstrated in human prostate cell lines that ETV4 overexpression have a role in cell proliferation [34], and this effect has been associated with the modulation of a set of cell cycleregulating genes
Summary
ETV4 is one of the ETS proteins overexpressed in prostate cancer (PC) as a result of recurrent chromosomal translocations. The results have not been always concordant: some studies suggest that ERG or ETV1 overexpression promotes premalignant in situ lesions (equivalent to prostatic intraepithelial neoplasia, PIN) [8,9,10,11,12], whereas other studies suggest that this overexpression is not sufficient to cause the onset of cancer [13,14,15,16,17,18]. These variable results may be related to many factors such as transgene expression levels, transgene integration site, transgene structure, and what promoter drives transgene expression. The genetic background of mice and the timing of the analysis may play a role, as in the case of human patients
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