A genome-wide RNAi screen identifies FOXO4 as a metastasis-suppressor through counteracting PI3K/AKT signal pathway in prostate cancer.
A genome-wide RNAi screen identifies FOXO4 as a metastasis-suppressor through counteracting PI3K/AKT signal pathway in prostate cancer.
复制标题
全基因组 RNAi 筛选将 FOXO4 鉴定为通过抵消前列腺癌 PI3K/AKT 信号通路的转移抑制剂
DOI：
10.1371/journal.pone.0101411
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Gelman IH
中科院分区：
文献类型：
--
作者：
Su B;Gao L;Baranowski C;Gillard B;Wang J;Ransom R;Ko HK;Gelman IH
PI3K/AKT信号通路的激活是去势-复发性前列腺癌(CR-CAP)的主要致死表型。在这里，我们使用基因组shRNA筛选确定了PI3K/AKT失活的下游靶点FOXO4，作为潜在的CAP转移抑制因子。FOXO4蛋白水平与一组人类帽子细胞系的侵袭潜能呈负相关，而mRNA水平的降低与临床转移发生率的增加相关。FOXO4在人LNCaP细胞中的敲除(KD)可导致体外侵袭增加和体内淋巴转移(LN)增加，而不影响增殖或凋亡指数。FOXO1的Kd增加了Matrigel侵袭力，而FOX03没有发现。比较FOXO4-KD在培养的LNCaP细胞、原发肿瘤和LN转移中的差异表达基因，发现一组上调的基因，包括PIP、CAMK2N1、PLA2G16和PGC，如果被siRNA敲除，可以降低与FOXO4缺乏相关的侵袭性增加。虽然这些基因中只有一部分编码FOXO启动子结合位点，但它们都是RUNX2诱导的，并且在FOXO4-KD细胞中，RUNX2与PIP启动子的结合增加。事实上，FOXO4的强制表达逆转了LNCaP/shFOXO4细胞侵袭性的增加；FOXO4的强制表达并没有改变RUNX2的蛋白水平，但它减少了RUNX2与PIP启动子的结合，导致PIP下调。最后，在人类CAP患者中，FOXO4，而不是FOXO1或FOX03，下调表达与降低无转移生存率之间存在相关性。我们的数据强烈表明，PI3K/AKT介导的CAP转移侵袭力的增加与FOXO4的缺失有关，而诱导FOXO4重新表达的机制可能会抑制CAP的转移侵袭力。
Activation of the PI3K/AKT signal pathway is a known driving force for the progression to castration-recurrent prostate cancer (CR-CaP), which constitutes the major lethal phenotype of CaP. Here, we identify using a genomic shRNA screen the PI3K/AKT-inactivating downstream target, FOXO4, as a potential CaP metastasis suppressor. FOXO4 protein levels inversely correlate with the invasive potential of a panel of human CaP cell lines, with decreased mRNA levels correlating with increased incidence of clinical metastasis. Knockdown (KD) of FOXO4 in human LNCaP cells causes increased invasion in vitro and lymph node (LN) metastasis in vivo without affecting indices of proliferation or apoptosis. Increased Matrigel invasiveness was found by KD of FOXO1 but not FOXO3. Comparison of differentially expressed genes affected by FOXO4-KD in LNCaP cells in culture, in primary tumors and in LN metastases identified a panel of upregulated genes, including PIP, CAMK2N1, PLA2G16 and PGC, which, if knocked down by siRNA, could decrease the increased invasiveness associated with FOXO4 deficiency. Although only some of these genes encode FOXO promoter binding sites, they are all RUNX2-inducible, and RUNX2 binding to the PIP promoter is increased in FOXO4-KD cells. Indeed, the forced expression of FOXO4 reversed the increased invasiveness of LNCaP/shFOXO4 cells; the forced expression of FOXO4 did not alter RUNX2 protein levels, yet it decreased RUNX2 binding to the PIP promoter, resulting in PIP downregulation. Finally, there was a correlation between FOXO4, but not FOXO1 or FOXO3, downregulation and decreased metastasis-free survival in human CaP patients. Our data strongly suggest that increased PI3K/AKT-mediated metastatic invasiveness in CaP is associated with FOXO4 loss, and that mechanisms to induce FOXO4 re-expression might suppress CaP metastatic aggressiveness.
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