An ultraconserved snoRNA-like element in long noncoding RNA CRNDE promotes ribosome biogenesis and cell proliferation (CRISPRi screen) [786-O]

Cancer cells frequently upregulate ribosome production to support tumorigenesis. While small nucleolar RNAs (snoRNAs) are critical for ribosome biogenesis, the roles of other classes of noncoding RNAs in this process remain largely unknown. Here we performed CRISPR screens to identify essential long noncoding RNAs (lncRNAs) in renal cell carcinoma (RCC) cells. This revealed that an alternatively-spliced isoform of lncRNA Colorectal Neoplasia Differentially Expressed containing an ultraconserved element (UCE), referred to as CRNDEUCE, is required for RCC cell proliferation. CRNDEUCE localizes to the nucleolus and promotes 60S ribosomal subunit biogenesis. The UCE of CRNDE functions as an unprocessed C/D box snoRNA that directly interacts with ribosomal RNA precursors. This facilitates delivery of eIF6, a key 60S biogenesis factor, which binds to CRNDEUCE through a sequence element adjacent to the UCE. These findings highlight the functional versatility of snoRNA sequences and expand the known mechanisms through which noncoding RNAs orchestrate ribosome biogenesis. A genome-wide CRISPRi screen was performed in 786-O cells expressing dCas9. Two independent clones were used for the screen. Samples were collected before (D0, control samples) and 15 days after prolonged growth (D15).

癌细胞通常会上调核糖体生成以支持肿瘤发生。尽管小核仁RNA（small nucleolar RNAs, snoRNAs）对核糖体生物发生至关重要，但其他类别非编码RNA在此过程中的作用仍未得到充分解析。本研究通过CRISPR筛选，鉴定了肾细胞癌（renal cell carcinoma, RCC）细胞中必需的长链非编码RNA（long noncoding RNAs, lncRNAs）。筛选结果显示，携带超保守元件（ultraconserved element, UCE）的长链非编码RNA结直肠肿瘤差异表达基因（Colorectal Neoplasia Differentially Expressed, CRNDE）的可变剪接异构体（被命名为CRNDEUCE）是肾细胞癌细胞增殖所必需的。CRNDEUCE定位于核仁，并可促进60S核糖体亚基的生物发生。CRNDE所携带的UCE可作为未加工的C/D框小核仁RNA，直接与核糖体RNA前体相互作用。这一过程能够促进关键60S生物发生因子eIF6的递送，而eIF6可通过UCE邻近的序列元件与CRNDEUCE结合。本研究结果揭示了小核仁RNA序列的功能多样性，并拓展了非编码RNA调控核糖体生物发生的已知机制。本研究在表达dCas9的786-O细胞中开展了全基因组CRISPR干扰（CRISPRi）筛选，实验使用两株独立克隆，并分别在增殖初始阶段（D0，对照样本）以及长期培养15天后（D15）收集样本。