NuRD复合体的新型亚基在促进胰腺癌细胞血管新生和转移中的作用机制研究

NuRD (The Nucleosome Remodelling and Histone Deacetylase complex) contains several core subunits and has also been implicated in the regulation of transcriptional events that are integral to cancer progression. There was only one paper that MTA1/NuRD is highly expressed in pancreatic cancer cell line. However, the mechanistic role of NuRD in pancreatic cancer cells remains an intriguing topic. In light of our previous finding that histone methyltransferase G9A is a component of the NuRD complex, we are interested in whether additional subunit or enzymatic activities that are associated with this complex and whether the role, if any, of NuRD is involved in the angiogenesis and metastasis of pancreatic cancer cells. We will focus on the search for histone methyltransferase SET8 as a new subunit of NuRD complex by a quantitative approach based on affinity purification-mass spectrometry combined with fast protein liquid chromatography system. Chromatin immunoprecipitation sequence will be used to uncover the molecular mechanisms for the target genes regulated by SET8 and NuRD. Molecular, biochemical, cell biological techniques as well as model animal strategy will be further used to reveal the biological significance for the complex. Over all, we will elaborate that SET8 is a new subunit of the NuRD complex, and involved in angiogenesis and metastasis of pancreatic cancer, providing a molecular mechanism underlying the role of SET8/NuRD in cancer metastasis and supporting the pursuit of SET8/NuRD as potential targets for pharmacological intervention in patients with pancreatic cancer.

NuRD是一个包含多亚基的复合体，其在多种恶性肿瘤的发生发展中发挥表观遗传学调控作用，但在胰腺癌中的作用尚不清楚。申请者前期研究证实，组蛋白甲基转移酶G9A（H3K9位点）可以作为NuRD复合体的新亚基而发挥生物学作用。此外，本项目申请者在预实验中通过亲和纯化-质谱、快速蛋白液相色谱等高通量方法筛选出组蛋白甲基转移酶SET8（H4K20位点）可能是胰腺癌细胞中NuRD复合体的新亚基。本项目将进一步明确SET8与NuRD复合体在胰腺癌细胞中的相互作用及其调控机制。利用免疫共沉淀测序，确定SET8和NuRD复合体在胰腺癌细胞中共同调控的下游靶基因。继而，在离体培养的胰腺癌细胞、胰腺癌种植瘤动物模型及胰腺癌患者的组织标本中，进一步探讨SET8作为NuRD复合体的新亚基对胰腺癌细胞的血管生成和转移的影响及作用机制。本项目将为研究胰腺癌发生发展的机制提供新的线索，并且为治疗胰腺癌提供潜在的作用靶点。

胰腺癌是一种恶性程度极高，病情发展快，迅速恶化，预后很差的消化系统肿瘤，既存在基因突变又存在表观遗传变化。NuRD复合体是一个包含多亚基的复合体，其在多种恶性肿瘤的发生发展中发挥表观遗传学调控作用，但在胰腺癌中的作用尚不清楚。本项目利用生物信息学分析发现，MTA2/NuRD复合体在胰腺癌组织中高表达，通过免疫组化染色的方法，检测MTA2与胰腺癌分级程度相关。通过染色质免疫共沉淀测序鉴定出肿瘤抑制因子PTEN是MTA2调控胰腺癌进程的靶基因，从细胞水平和动物水平两个层面验证MTA2通过抑制PTEN，激活PI3K/Akt信号通路，促进胰腺癌的增殖，迁移和侵润等作用。鉴于组蛋白甲基转移酶SET8在上皮间质化，细胞增殖，DNA损伤修复中的重要作用，推测SET8可能在肿瘤的发生发展过程中发挥重要作用。那么，对于胰腺癌来说，SET8发挥着怎样的功能呢，机制尚不明确。本研究证明，在胰腺癌细胞中，证明SET8作为MTA1/NuRD复合体的亚基、发挥酶活性，协同MTA1/NuRD复合体在胰腺癌细胞中发挥转录调控作用的机制。本研究将为研究胰腺癌发生发展的机制提供新的线索，并且为治疗胰腺癌的治疗提供潜在的作用靶点。