The CtBP1–p300–FOXO3a transcriptional complex represses the expression of the apoptotic regulators Bax and Bim in human osteosarcoma cells

Abstract

C-terminal binding protein 1 (CtBP1), a well-known transcriptional corepressor, functions as an oncogene in multiple cancer types, including osteosarcoma, by modulating the transcription of many tumor suppressors, such as cadherin 1 (CDH1), phosphatase and tensin homolog (PTEN), Bcl2-associated X (Bax), Bcl-2-interacting mediator (Bim), and cyclin-dependent kinase inhibitor 1A (CDKN1A). However, it is still unclear how CtBP1 regulates the expression of these downstream targets. Here, we identified that CtBP1 is overexpressed in osteosarcoma cells and found that CtBP1 directly interacts with the transcription factor forkhead box O3 (FOXO3a) and the histone acetyltransferase p300 in vivo and in vitro. Through microarray analysis, we found that CtBP1 negatively regulates FOXO3a levels. In contrast to the CtBP1 level, the FOXO3a expression level was found to be significantly reduced in osteosarcoma cells. Knockdown of CtBP1 or overexpression of FOXO3a in U2OS cells resulted in different gene expression patterns, and the former caused upregulation of CtBP1 downstream target genes such as CDH1, PTEN, Bax, Bim, and CDKN1A, whereas the latter caused upregulation of Bax and Bim, but not CDH1, PTEN, and CDKN1A. Further analysis indicated that the CtBP1-p300-FOXO3a transcriptional complex specifically binds to the promoters of Bax and Bim. Inhibition of CtBP1 by the constitutive expression of Pep1-E1A WT peptide in U2OS and OSA cells reversed oncogenic phenotypes, including colony formation, cellular proliferation, and migration, and limited tumor growth in vivo. Together our results demonstrated that the CtBP1-p300-FOXO3a transcriptional complex represses the expression of the apoptotic regulators Bax and Bim in human osteosarcoma cells and that targeting CtBP1-mediated transcriptional events might be a potential therapeutic strategy for the osteosarcoma treatment.

Transforming growth factor beta (TGF-β) is activated by the CtBP2-p300-AP1 transcriptional complex in chronic renal failure

“…Comparing these two protein lists, we found that p300 and CtBP2 overlapped (Supplementary Table 4 and 5). This result, together with previous publications [23,30,31], suggested that these two proteins had high possibilities of being recruited by c-Jun and c-FOS, thereby forming two transcriptional complexes. To verify this hypothesis, we primarily examined the in vivo association of CtBP2, p300 and AP-1 subunits.…”

“…Cells were grown in Dulbecco's modified Eagle's medium (DMEM)/F12 medium (Thermo Fisher Scientific, Waltham, MA, USA, #12634010) containing 10% fetal bovine serum (FBS) (Sigma-Aldrich, Shanghai, China, #F4135) and 1% penicillin-streptomycin antibiotics (Sigma-Aldrich, #P4333) at 37 °C. Cells under 80% confluences were used for transfection to knock down or overexpress genes following a previous protocol [30]. Briefly, siRNAs specifically targeting genes were purchased from Santa Cruz Biotechnology (Dallas, TX, USA), and their catalog numbers were sc-29410 (p65), sc-29407 (p50), sc-29223 (c-Jun), sc-29221 (c-FOS) and sc-37767 (CtBP2).…”

“…The purified Flag-c-Jun and Flag-c-FOSassociated complexes were separated on an SDS gel and stained with a Pierce Silver Stain Kit (ThermoFisher Scientific, #24612). The stained bands were excised and fragmented as described previously [30]. The resulting proteins were analyzed with a Q Exactive TM HF-X Hybrid Quadrupole-Orbitrap TM Mass Spectrometer (ThermoFisher Scientific, #0726042).…”

p300 is upregulated by docetaxel and is a target in chemoresistant prostate cancer

“…It has been described that the cysteine protease USP24 stabilizes p300 and thereby increases acetylation of histone h3 (Wang et al 2018). The CtBP1-p300-FOXO 3a complex was found to repress apoptotic regulators Bax and Bim in osteosarcoma cells (Li et al 2019). Cell proliferation is regulated by p300 in complex with the transcriptional repressor YY1 and HDAC2 (Tang et al 2019).…”

The Monocyte-Derived Exosomal CLMAT3 Activates the CtBP2-p300-NF-κB Transcriptional Complex to Induce Proinflammatory Cytokines in ALI

“…Several studies have shown that CtBP1/2 (C-terminal binding proteins 1 and 2) can interact with p300 and HDACs (histone deacetylases) through the PXDLS motif. 29 , 30 , 31 The current evidence suggests that CtBPs have both transrepression and transactivation roles in regulating the expression of genes, miRNAs, and lncRNAs. 32 , 33 , 34 , 35 Generally, CtBPs interact with p300, HDACs, or transcription factors such as ZEB1 (zinc finger E-box-binding homeobox 1) and FOXP2 (Forkhead box protein P2).…”