前列腺癌中废除KLF5的乙酰化修饰将其由抑癌基因转变为促癌基因

发布时间：2018-04-29 11:55

本文选题：KLF5 + TGF-β　；参考：《南开大学》2014年博士论文

【摘要】：前列腺癌是男性泌尿系统最常见的恶性肿瘤,并且容易发生浸润和迁移。在不同的国家和地区,前列腺癌的发病率存在差异,在欧美发达国家和地区,前列腺癌具有非常高的发病率,也是男性恶性肿瘤主要致死原因之一；在我国,前列腺癌的发病率和致死率低于欧美发达国家,但是随着近年来人口老龄化的加剧以及饮食习惯的改变,我国男性前列腺癌的发病率呈现不断上升的趋势。为了提高前列腺癌的治疗效果以及患者的生活质量,许多前列腺癌相关基因的功能及分子机制有待研究。作为基础转录因子,KLF5(Kruppel-like factor5)同时具有肿瘤抑制功能和肿瘤促进功能,尽管这种功能转换的机制尚不明确。我们以前的研究结果表明,在正常上皮细胞中,KLF5促进细胞增殖,而经过TGF-p处理之后,KLF5抑制细胞增殖,这是因为TGF-β可以招募p300诱导KLF5的乙酰化,进而改变KLF5转录复合体以及下游靶基因p15和MYC的表达,从而将KLF5的功能由促进细胞增殖转变为抑制细胞增殖。在这篇文章中,我们检测了KLF5的乙酰化修饰形式是否决定了其在肿瘤发生过程中的相反作用。首先,我们选用前列腺癌细胞系DU145和PC-3为细胞模型,这两种细胞的均低表达KLF5,并且它们的增殖均受到TGF-β的抑制。在这两种细胞系中,我们通过病毒感染的方法构建稳定表达野生型KLF5以及KLF5突变体K369R的克隆,K369R突变体可以组成型废除KLF5的乙酰化修饰。体外细胞培养的研究结果表明,野生型KLF5阻抑细胞增殖,抑制MYC表达,促进p15表达,而K369R突变体丧失阻抑细胞增殖的功能,促进MYC表达。这些研究结果提示我们,KLF5抑制细胞增殖的功能依赖于其自身的乙酰化修饰。接下来,我们用上述稳定克隆进行动物实验,同样的,KLF5在乙酰化的状态下抑制肿瘤生长,而去乙酰化的K369R突变体可以将KLF5的功能转变为促进肿瘤生长。随后,我们在裸鼠体内注射TGF-β抑制剂SB431542,结果表明,封闭TGF-p信号通路可以废除KLF5的抑癌功能。最后,我们将空载对照pLHCX、野生型KLF5以及K369R突变体的成瘤样本送RNA-Seq检测,结果表明,AcKLF5和unAcKLF5分别调控不同的下游靶基因,以不同的方式影响不同的生物学过程,从而决定二者在肿瘤发生发展过程中的相反功能。综上所述,KLF5的乙酰化形式可以通过调控MYC和p15以及相关下游基因表达的方式抑制前列腺癌的细胞增殖和肿瘤生长,废除KLF5的乙酰化修饰将KLF5的抑癌功能转变为促癌功能。这些结果为研究KLF5由抑癌功能转变为促癌功能的机制提供了新的视野,同时也支持cKLF5和inAcKLF5在TGF-β的抑癌功能和促癌功能的转换中发挥作用。
[Abstract]:Prostate cancer is the most common malignant tumor in the male urinary system and is prone to invasion and migration. In different countries and regions, the incidence of prostate cancer is different. In developed countries and regions in Europe and America, prostate cancer has a very high incidence, which is also one of the main causes of death of male malignant tumors. The morbidity and fatality rate of prostate cancer is lower than that of developed countries in Europe and America. However, with the aggravation of population aging and the change of dietary habits in recent years, the incidence of prostate cancer in men in China is on the rise. In order to improve the therapeutic effect of prostate cancer and the quality of life of patients, the function and molecular mechanism of many prostate cancer related genes need to be studied. As a basic transcription factor, KLF5 / Kruppel-like factor5 has both tumor suppressor function and tumor-promoting function, although the mechanism of this functional transformation is unclear. Our previous studies have shown that KLF5 promotes cell proliferation in normal epithelial cells, whereas KLF5 inhibits cell proliferation after TGF-p treatment, because TGF- 尾 can recruit p300-induced acetylation of KLF5. Furthermore, the expression of KLF5 transcription complex and the downstream target genes p15 and MYC were altered, and the function of KLF5 was changed from promoting cell proliferation to inhibiting cell proliferation. In this paper, we examined whether the acetylation modification of KLF5 determines its opposite role in tumorigenesis. Firstly, the prostate cancer cell lines DU145 and PC-3 were used as cell models, both of which were low expressed KLF5, and their proliferation was inhibited by TGF- 尾. In these two cell lines, we constructed the stable expression of wild-type KLF5 by virus infection and the clone K369R of KLF5 mutant K369R, which can constitutively abolish the acetylation modification of KLF5. The results of cell culture in vitro showed that wild-type KLF5 inhibited cell proliferation, inhibited MYC expression and promoted p15 expression, while K369R mutant lost the function of inhibiting cell proliferation and promoting MYC expression. These results suggest that the inhibitory effect of KLF5 on cell proliferation depends on its own acetylation modification. Next, we used the above stable clones in animal experiments. The same KLF5 inhibited tumor growth in acetylated state, while deacetylated K369R mutant could transform the function of KLF5 into promoting tumor growth. Subsequently, we injected TGF- 尾 inhibitor SB431542 into nude mice. The results showed that blocking the TGF-p signaling pathway could abolish the anti-cancer function of KLF5. Finally, we sent non-loaded control pLHCX, wild-type KLF5 and K369R mutant for RNA-Seq detection. The results showed that AcKLF5 and unAcKLF5 regulated different downstream target genes and affected different biological processes in different ways. This determines the opposite function of both in the tumor development process. In conclusion, the acetylation of KLF5 can inhibit the proliferation and tumor growth of prostate cancer cells by regulating the expression of MYC, p15 and related downstream genes, and abolish the acetylation modification of KLF5 to transform the tumor suppressor function of KLF5 into carcinogenic promotion function. These results provide a new perspective for the study of the mechanism of KLF5 transforming from tumor suppressor function to promoting cancer function, and also support the role of cKLF5 and inAcKLF5 in the transformation of tumor suppressor function and carcinogenic function of TGF- 尾.
【学位授予单位】：南开大学
【学位级别】：博士
【学位授予年份】：2014
【分类号】：R737.25

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