Kim Taewan, Jeon Young-Jun, Cui Ri, Lee Ji-Hoon, Peng Yong, Kim Sung-Hak, Tili Esmerina, Alder Hansjuerg, Croce Carlo M
Department of Molecular and Cellular Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX (TK); Department of Molecular Virology, Immunology and Medical Genetics (TK, YJJ, RC, ET, HA, CMC), Department of Neurological Surgery (SHK), and Department of Anesthesiology (ET), Wexner Medical Center, The Ohio State University, OH; School of Biological Sciences, Seoul National University and National Creative Research Initiative Center for Symbiosystem, Seoul, Republic of Korea (JHL); State Key Laboratory of Biotherapy/Collaborative Innovation Center of Biotherapy, West China Hospital, Sichuan University, Chengdu, China (YP); School of Life Sciences and Biotechnology, Korea University, Republic of Korea (SHK).
J Natl Cancer Inst. 2015 Feb 6;107(4). doi: 10.1093/jnci/dju505. Print 2015 Apr.
The functions of long noncoding RNAs (lncRNAs) have been identified in several cancers, but the roles of lncRNAs in colorectal cancer (CRC) are less well understood. The transcription factor MYC is known to regulate lncRNAs and has been implicated in cancer cell proliferation and tumorigenesis.
CRC cells and tissues were profiled to identify lncRNAs differentially expressed in CRC, from which we further selected MYC-regulated lncRNAs. We used luciferase promoter assay, ChIP, RNA pull-down assay, deletion mapping assay, LC-MS/MS and RNA immunoprecipitation to determine the mechanisms of MYC regulation of lncRNAs. Moreover, soft agar assay and in vivo xenograft experiments (four athymic nude mice per group) provided evidence of MYC-regulated lncRNAs in cancer cell transformation and tumorigenesis. The Kaplan-Meier method was used for survival analyses. All statistical tests were two-sided.
We identified lncRNAs differentially expressed in CRC (P < .05, greater than two-fold) and verified four lncRNAs upregulated and two downregulated in CRC cells and tissues. We further identified MYC-regulated lncRNAs, named MYCLos. The MYC-regulated MYCLos may function in cell proliferation and cell cycle by regulating MYC target genes such as CDKN1A (p21) and CDKN2B (p15), suggesting new regulatory mechanisms of MYC-repressed target genes through lncRNAs. RNA binding proteins including HuR and hnRNPK are involved in the function of MYCLos by interacting with MYCLo-1 and MYCLo-2, respectively. Knockdown experiments also showed that MYCLo-2, differentially expressed not only in CRC but also in prostate cancer, has a role in cancer transformation and tumorigenesis.
Our results provide novel regulatory mechanisms in MYC function through lncRNAs and new potential lncRNA targets of CRC.
长链非编码RNA（lncRNA）的功能已在多种癌症中得到鉴定，但lncRNA在结直肠癌（CRC）中的作用尚不清楚。已知转录因子MYC可调节lncRNA，并与癌细胞增殖和肿瘤发生有关。
对CRC细胞和组织进行分析，以鉴定在CRC中差异表达的lncRNA，从中进一步选择受MYC调节的lncRNA。我们使用荧光素酶启动子分析、染色质免疫沉淀、RNA下拉分析、缺失定位分析、液相色谱-串联质谱和RNA免疫沉淀来确定MYC调节lncRNA的机制。此外，软琼脂分析和体内异种移植实验（每组4只无胸腺裸鼠）提供了受MYC调节的lncRNA在癌细胞转化和肿瘤发生中的证据。采用Kaplan-Meier法进行生存分析。所有统计检验均为双侧检验。
我们鉴定了在CRC中差异表达的lncRNA（P <.05，大于两倍），并验证了在CRC细胞和组织中上调的4种lncRNA和下调的2种lncRNA。我们进一步鉴定了受MYC调节的lncRNA，命名为MYCLos。受MYC调节的MYCLos可能通过调节MYC靶基因如CDKN1A（p21）和CDKN2B（p15）在细胞增殖和细胞周期中发挥作用，提示MYC通过lncRNA抑制靶基因的新调控机制。包括HuR和hnRNPK在内的RNA结合蛋白分别通过与MYCLo-1和MYCLo-2相互作用参与MYCLos的功能。敲低实验还表明，不仅在CRC中而且在前列腺癌中差异表达的MYCLo-2在癌症转化和肿瘤发生中起作用。
我们的结果提供了通过lncRNA调节MYC功能的新调控机制以及CRC新的潜在lncRNA靶点。
