肿瘤干细胞（CSC）是肿瘤复发和转移的根源，诱导维持CSC多潜能“干性”的分子机制是CSC研究的核心问题。近期研究发现pre-RCs蛋白通过表观遗传调控参与诱导细胞多潜能“干性”，但具体机制还不清楚。我们前期结果证实，Cdc6、Mcm7在CSC恶性转化过程中显著上调，并分别结合于E-cadherin、INK4A/APF和EZH2基因启动子区。本项目拟①通过研究Cdc6-Bmi1-EZH2轴，阐明Cdc6协同Bmi1、EZH2调控E-cadherin、INK4a/ARF转录机制；②通过研究Mcm7-Sp1轴，阐明Mcm7协同Sp1调控EZH2转录机制；③通过研究PTEN-E2F及ZBTB4-Sp1-EZH2轴，阐明Mcm7内含miR-106b-25对Cdc6、Mcm7的促进作用。本项目的完成将从分子水平建立DNA复制与转录之间的联系，深入揭示pre-RCs蛋白促进CSC恶性转化的分子机制。

Cancer stem cells (CSC) are subpopulation cells that have an unlimited proliferative potential. CSC are resistant to conventional therapies, which cause tumor recurrence and remote metastasis. Thus, CSC-targeting approach appears as a major goal toward the improvement of the clinical outcome of patients with incurable tumors. .An essential issue of the anti-CSC research is the molecular mechanism underlying the acquirement and maintenance of pluripotency. Recently, the pre-RC proteins, which are essential for DNA replication, were shown to have ability to regulate gene transcription, therefore contribute to epigenetic regulation. Our previous results show that pre-RC proteins Cdc6 and Mcm7 were significantly up-regulated during the CSC malignant transformation. Furthermore, Cdc6 is found to located in the promotor region of E-cadherin and INK4A/APF gene, and Mcm7 locate in the promotor region of polycomb protein EZH2 gene. Thus, we hypotheses that in the CSC malignant transformation, Cdc6 may cooperated with Bmi1 and EZH2 to silence E-cadherin and INK4A/APF gene, and Mcm7 may cooperated with EZH2 to enhance EZH2 gene. Cancer stem cells (CSC) are subpopulation cells that have an unlimited proliferative potential. CSC are resistant to conventional therapies, which cause tumor recurrence and remote metastasis. Thus, CSC-targeting approach appears as a major goal toward the improvement of the clinical outcome of patients with incurable tumors. .An essential issue of the anti-CSC research is the molecular mechanism underlying the acquirement and maintenance of pluripotency. Recently, the pre-RC proteins, which are essential for DNA replication, were shown to have ability to regulate gene transcription, therefore contribute to epigenetic regulation. Our previous results show that pre-RC proteins Cdc6 and Mcm7 were significantly up-regulated during the CSC malignant transformation. Furthermore, Cdc6 is found to located in the promotor region of E-cadherin and INK4A/APF gene, and Mcm7 locate in the promotor region of polycomb protein EZH2 gene. Thus, we hypotheses that in the CSC malignant transformation, Cdc6 may cooperated with Bmi1 and EZH2 to silence E-cadherin and INK4A/APF gene, and Mcm7 may cooperated with EZH2 to enhance EZH2 gene. Moreover, the miR-106b-25, a Mcm7 intronic miRNA, may exert epigenetic regulation to further enhance the oncogenic activity of Cdc6 and Mcm7..In this project, we plan to test our hypothesis by pursuing the following three specific aims: (1) to clarify whether Cdc6, Bmi1 and EZH2 synergistically control E‑cadherin and INK4a/ARF gene transcription. We will investigate the Cdc6-Bmi1-EZH2 axis in the CSC to test the hypothesis that Cdc6 collaborate with Bmi1 and EZH2 to repress E‑cadherin and INK4a/ARF gene transcription. (2) To clarify the mechanism by which Mcm7 regulate EZH2 gene transcription. We will investigate the Mcm7-Sp1 axis to test the hypothesis that Mcm7 collaborate with Sp1 to promote EZH2 gene transcription. (3) To Investigate the PTEN-E2F and ZBTB4-Sp1-EZH2 axis to clarify the promoted effect of miR-106b-25 on Cdc6 and Mcm7.