Department of Biotechnology, Pondicherry Central University, Pondicherry, 605014, India.
Med Oncol. 2019 Jan 21;36(2):20. doi: 10.1007/s12032-018-1240-2.
CUL4A; an E3 ubiquitin ligase is involved in the degradation of negative regulators of cell cycle such as p21, p27, p53, etc., through polyubiquitination-mediated protein degradation. The functional role(s) of CUL4A proteins on their targets are well characterized; however, the transcriptional regulation of CUL4A, particularly at its promoter level is not yet studied. Therefore, in this study, using computational tools, we found cAMP responsive elements (CRE) at the locations of - 926 and - 764 with respect to transcription state site + 1 of CUL4A promoter. Hence, we investigated the role of CREB on the regulation of CUL4A transcription. Our chromatin immunoprecipitation (ChIP) data clearly showed increased levels of promoter occupancy of both CREB and pCREB on both CREs of CUL4A promoter. As expected, the expression of CUL4A increases and decreases upon the overexpression of and knocking down of CREB, respectively. Moreover, the inhibition of ERK pathway by U0126 not only reduces the CREB activation but also the CUL4A levels suggesting that CREB is the upstream activator of CUL4A transcription. The reduction of CUL4A levels upon the knocking down of CREB or by U0126 treatment increases the protein levels of CUL4A substrates such as p21 and p27. It is reported that CUL4A activates the ERK1/2 transcription and ERK1/2 pathway activates the CREB by phosphorylation. Based on our data and earlier findings, we report that CREB regulates the CUL4A levels positively which in turn activates the CREB through ERK1/2 pathway in the form of auto-regulatory looped mechanism.This suggests that CUL4A might be involved in proliferation of cancer cells by regulating the ERK1/2 and CREB signaling.
CUL4A 是一种 E3 泛素连接酶，通过多泛素化介导的蛋白质降解参与细胞周期负调控因子如 p21、p27、p53 等的降解。CUL4A 蛋白对其靶标的功能作用已得到很好的描述；然而，CUL4A 的转录调控，特别是在启动子水平上尚未得到研究。因此，在这项研究中，我们使用计算工具在 CUL4A 启动子的转录起始位点+1 处的-926 和-764 位置发现了 cAMP 反应元件（CRE）。因此，我们研究了 CREB 对 CUL4A 转录的调节作用。我们的染色质免疫沉淀（ChIP）数据清楚地显示，在 CUL4A 启动子的两个 CRE 上，CREB 和 pCREB 的启动子占有率都增加了。正如预期的那样，CREB 的过表达会增加 CUL4A 的表达，而 CREB 的敲低则会降低 CUL4A 的表达。此外，U0126 抑制 ERK 通路不仅降低了 CREB 的激活，也降低了 CUL4A 的水平，这表明 CREB 是 CUL4A 转录的上游激活子。CREB 的敲低或 U0126 处理导致 CUL4A 水平降低，从而增加了 CUL4A 底物如 p21 和 p27 的蛋白水平。据报道，CUL4A 激活 ERK1/2 的转录，而 ERK1/2 途径通过磷酸化激活 CREB。基于我们的数据和早期的发现，我们报告说 CREB 正向调节 CUL4A 的水平，CUL4A 反过来又通过 ERK1/2 途径以自动调节循环机制激活 CREB。这表明 CUL4A 可能通过调节 ERK1/2 和 CREB 信号参与癌细胞的增殖。
