Abstract B031: CREB1 and ATF1 differentially regulate terminal prostate luminal differentiation by controlling the timing of ING4 expression, while CREB1 prevents ING4 expression upon PTEN loss in prostate cancer
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Abstract B031: CREB1 and ATF1 differentially regulate terminal prostate luminal differentiation by controlling the timing of ING4 expression, while CREB1 prevents ING4 expression upon PTEN loss in prostate cancer
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<jats:title>Abstract</jats:title>
<jats:p>Many genes aberrantly expressed in prostate cancer are involved in normal basal to luminal cell differentiation. We previously demonstrated that transient ING4 expression is required for luminal cell differentiation and is downregulated in ~60% of primary prostate tumors. We further demonstrated in a primary prostate cancer model overexpressing ERG, MYC, and shPTEN (EMP) that loss of PTEN was responsible for ING4 loss. Furthermore, half of the human tumor samples that lose ING4 have also lost PTEN. However, we did not know how PTEN loss inhibits ING4 expression. Utilizing our in vitro differentiation model, whereby prostate basal epithelial cells (iPrEC) treated with KGF and androgen induce a suprabasal layer of luminal-like cells, and RNA-seq we identified transcriptional nodes required for luminal differentiation. Differentially expressed genes were analyzed by GeneGo to identify enriched transcription-factor signatures. Of the ~600 differentially regulated genes during differentiation, the largest signature (29% of genes) was CREB/ATF targets. Induction of Blimp1, Claudin1, and Plk2 and inhibition of Chek1 were further validated by qRT-PCR and immunoblotting.</jats:p>
<jats:p>CREB/ATF bind constitutively to open chromatin CRE elements in the promoters of genes and are activated through signaling-induced phosphorylation at Ser133 by kinases, including AKT. We found that both CREB1 and ATF1 are inducibly phosphorylated midway through luminal differentiation, with ATF1 preceding CREB1. Knockdown of CREB1 with shRNA increased ING4, accelerated differentiation, and induced premature luminal cell death. Conversely, knockdown of ATF1 blocked ING4 induction and prevented suprabasal layer formation. CREB1/ATF1 ChIP was enriched at the ING4 promoter at mid-differentiation, when ING4 expression peaks. Additionally, CREB1/ATF1 was constitutively bound to the promoter of JFK, an E3-ligase that targets ING4 and whose mRNA levels increase during differentiation. Thus, we propose that ATF1 is required to induce ING4 transcription, while CREB1 suppresses ING4 and simultaneously activates its E3-ligase to tightly control the timing of ING4 expression. We compared the gene signature of luminal cells to that of the tumorigenic EMP cells and, surprisingly, found that 30% of the differentially expressed genes were also CREB/ATF targets. However, there is less than 10% overlap in these targets, indicating that CREB/ATF control distinct subsets of genes in differentiated luminal cells versus cancer cells. Some of EMP-specific CREB/ATF targets included GATA2, TWIST1, Necdin, and PPM1F, which were further validated by qRT-PCR and immunoblotting. CREB1 and ATF1 were highly phosphorylated in EMP cells and knockdown of CREB1 restored ING4 expression and suprabasal formation.</jats:p>
<jats:p>Our working model is that AKT activation upon PTEN loss in transiently differentiating luminal cells results in premature and constitutive activation of CREB1/ATF1 bound to genes prior to induction of the ING4 chromatin switch. This prevents ING4 induction and the chromatin rearrangements required for terminal differentiation. In normal PrECs, CREB/ATF1 activation is tightly controlled by as yet undetermined factors and is only permitted when the proper CRE binding sites are exposed. This model helps to explain how loss of PTEN disrupts luminal cell terminal differentiation to promote prostate cancer oncogenesis.</jats:p>
<jats:p>Citation Format: McLane Watson, Penny Berger, Sander Frank, Mary Winn, Cindy Miranti. CREB1 and ATF1 differentially regulate terminal prostate luminal differentiation by controlling the timing of ING4 expression, while CREB1 prevents ING4 expression upon PTEN loss in prostate cancer [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr B031.</jats:p>