Alternative polyadenylation (APA) has emerged as a prevalent feature associated with cancer development and progression. The advantage of APA to tumor progression is to induce oncogenes through 3′-UTR shortening, and to inactivate tumor suppressor genes via the re-routing of microRNA competition. We previously identified the Mammalian Cleavage Factor I-25 (CFIm25) (encoded by Nudt21 gene) as a master APA regulator whose expression levels directly impact the tumorigenicity of glioblastoma (GBM) in vitro and in vivo. Despite its importance, the role of Nudt21 in GBM development is not known and the genes subject to Nudt21 APA regulation that contribute to GBM progression have not been identified. Here, we find that Nudt21 is reduced in low grade glioma (LGG) and all four subtypes of high grade glioma (GBM). Reduced expression of Nudt21 associates with worse survival in TCGA LGG cohorts and two TCGA GBM cohorts. Moreover, although CFIm25 was initially identified as biochemically associated with both CFIm59 and CFIm68, we observed three CFIm distinct subcomplexes exist and CFIm59 protein level is dependent on Nudt21 expression in GBM cells, but CFIm68 is not, and that only CFIm59 predicts prognosis of GBM patients similar to Nudt21. Through the use of Poly(A)-Click-Seq to characterize APA, we define the mRNAs subject to 3′-UTR shortening upon Nudt21 depletion in GBM cells and observed enrichment in genes important in the Ras signaling pathway, including Pak1. Remarkably, we find that Pak1 expression is regulated by Nudt21 through its 3′-UTR APA, and the combination of Pak1 and Nudt21 expression generates an even stronger prognostic indicator of GBM survival versus either value used alone. Collectively, our data uncover Nudt21 and its downstream target Pak1 as a potential “combination biomarker” for predicting prognosis of GBM patients.

替代性聚腺苷酸化（APA）已成为与癌症发展和进展相关的普遍特征。APA对肿瘤进展的优势是通过缩短3'-UTR诱导癌基因，并通过重新路由microRNA竞争使肿瘤抑制基因失活。我们以前鉴定了哺乳动物卵裂因子I-25（CFIm25）（由Nudt21基因编码）作为APA主调节剂，其表达水平直接影响体外和体内胶质母细胞瘤（GBM）的致瘤性。尽管其重要性，Nudt21在GBM发育中的作用尚不清楚，并且尚未鉴定出受Nudt21 APA调控的基因有助于GBM的进展。在这里，我们发现Nudt21低度神经胶质瘤（LGG）和高度神经胶质瘤（GBM）的所有四个亚型均降低。Nudt21的表达降低与TCGA LGG队列和两个TCGA GBM队列中较差的生存率相关。此外，尽管最初将CFIm25与CFIm59和CFIm68进行了生化相关联，但我们观察到存在三个CFIm不同的亚复合物，并且CFIm59蛋白的水平取决于GBM细胞中Nudt21的表达，但CFIm68并非如此，只有CFIm59可以预测GBM患者的预后类似于Nudt21。通过使用Poly（A）-Click-Seq表征APA，我们定义了在GBM细胞中Nudt21耗竭后3'-UTR缩短的mRNA，并观察到了重要的基因富集。Ras信号传导途径，包括Pak1。值得注意的是，我们发现Pak1表达受Nudt21通过其3'-UTR APA的调节，并且Pak1和Nudt21表达的组合产生的GBM生存率甚至比单独使用任一个值都更强。总体而言，我们的数据揭示了Nudt21及其下游靶标Pak1作为预测GBM患者预后的潜在“组合生物标志物”。