BCCIP binds to and activates its promoter in a YY1‐dependent fashion in HCT116 cells

Abstract

The restriction of Yin Yang 1 (YY1) at BRCA2 and CDKN1A/p21-interacting protein (BCCIP) transcriptional start site (TSS) proximal region in several human cancer cell lines was found by analyzation of ChIP-Seq database from UCSC Genome Browser (http://genome.ucsc.edu). However, whether the stabilization of YY1 by BCCIP impacts its recruitment in the BCCIP promoter region is unclear. Here, we present evidence that transcriptional regulation of YY1 on BCCIP is closely related to YY1 stability in HCT116 human colon cancer cells. YY1 stabilization was in turn regulated by BCCIP, suggesting the existence of a BCCIP-YY1 feedback loop in regulating BCCIP transcription by the YY1. Overexpression of BCCIP stabilized YY1 while knockdown of BCCIP reduced YY1 protein level. In addition, direct interaction between YY1 and BCCIP was confirmed by coimmunoprecipitation approach. Also, the N-terminus region of BCCIP, including the internal conserved domain (ICD), was responsible for binding with the amino acid 146-270 of YY1. More importantly, YY1 stability was related to the BCCIP/ICD domain-mediated YY1 ubiquitination pathway. Moreover, a limited BCCIP promoter region containing YY1 binding site (CCGCCATC) was tightly associated with the pGL4-BCCIP-Luc luciferase activity. In ChIP assays, shBCCIP lentiviral-mediated YY1 instability decreased recruitment of the YY1 at BCCIP TSS proximal region, which could not be restored by YY1 overexpression. Furthermore, knockdown of YY1 inhibited the binding of BCCIP itself at BCCIP promoter region proximal to TSS, demonstrating that transcriptional regulation of the YY1 on BCCIP can be modulated by BCCIP itself in a YY1-dependent fashion.

YY1/BCCIP Coordinately Regulates P53-Responsive Element (p53RE)-Mediated Transactivation of p21Waf1/Cip1

“…However, this induction was blocked in the pLVX-shBCCIP-transfected cells (Figure 6B, lane 4). In addition, consistent with previous reports [17], YY1 protein levels significantly declined in BCCIP knockdown cells with or without 5FU treatment (Figure 6B, lane 2/IB:YY1). Next, the coordinative effects of BCCIP/YY1 on p53 and p21 protein levels were examined by western blot in 250 µM 5FU-exposed HCT116 cells.…”

“…This view is also confirmed by later experiments showing that YY1 and INO80 together transcriptionally regulate BCCIP in HCT16 cells [18]. However, transcriptional regulation of YY1 on BCCIP is in turn modulated by BCCIP itself in a YY1-dependent fashion [17]. On the other hand, stably expressing shBCCIP abrogates the transactivation activity of p53 and results in low expression of p21 , suggesting the involvement of BCCIP in p21 transcription [19,20].…”

“…On the other hand, according to our previous reports, BCCIP can also stabilize intracellular YY1 protein [17]. So, to know whether the stabilization effect of BCCIP on p53 is impacted by YY1, the effect of YY1 on p53 was first examined.…”

“…No matter which sequence region was targeted, compared to shNT groups, knocking down BCCIP appeared to result in no big changes in p53RE luciferase activity reduction due to YY1. Based on our previous report [17], the 146–270 amino acids of YY1 are the binding region for BCCIP. We therefore designed experiments as shown in Figure 5D.…”

“…Recent research data demonstrates that BCCIP maintains YY1 protein stability by directly binding to it in HCT116 cells. Co-transfection/coimmunoprecipitation (CoIP) experiments have confirmed that YY1/146-270 amino acids are the binding region for BCCIP, and at the same time, the BCCIP/ICD domain plays a key role in regulating YY1 stability through the ubiquitin-proteasome-mediated degradation pathway [17]. Based on a chromatin immunoprecipitation (ChIP)-Seq database search from the University of California Santa Cruz (UCSC) Genome Browser (), the enrichment of YY1 at the BCCIP transcriptional start site (TSS) proximal region in several cancer cells including A549 lung cancer cells, HepG2 human hepatocellular carcinoma cells, and HCT116 human colon cancer cells was found, suggesting the function of YY1 in regulating BCCIP gene transcription.…”