LIN28B alters ribosomal dynamics to promote metastasis in MYCN-driven malignancy (RIP-Seq). LIN28B alters ribosomal dynamics to promote metastasis in MYCN-driven malignancy (RIP-Seq)

High expression of LIN28B is associated with aggressive malignancy and poor survival. Here, probing MYCN-amplified neuroblastoma as a model system, we show that LIN28B expression is associated with enhanced cell migration in vitro and invasive and metastatic behavior in murine xenografts. Sequence analysis of the polyribosome fraction of LIN28B-expressing neuroblastoma cells revealed let-7 independent enrichment of transcripts encoding components of the translational and ribosomal apparatus, particularly those with higher adenine/uridine (AU)-content, and depletion of transcripts of neuronal developmental programs. We further show that LIN28B utilizes both its cold shock and zinc finger RNA binding domains to preferentially interact with MYCN-induced transcripts of the ribosomal complex, enhancing their translation. These data demonstrate that LIN28B couples the MYCN-driven transcriptional program to enhanced ribosomal translation, thereby implicating LIN28B as a post-transcriptional driver of the metastatic phenotype. Overall design: Comparison of LIN28B WT BE2C versus LIN28B KO BE2C cells as well as re-expression of LIN28B WT, LIN28B MUT (full mutant of cold shock domain (CSD) and zinc knuckle domain) and single Mutants LIN28B CSD MUT/ZKD MUT upon LIN28B KO BE2C.

LIN28B高表达与侵袭性恶性表型及不良生存预后密切相关。本研究以MYCN扩增型神经母细胞瘤作为模型系统，证实LIN28B表达与体外细胞迁移能力增强、小鼠异种移植瘤的侵袭及转移行为呈正相关。对表达LIN28B的神经母细胞瘤细胞的多聚核糖体组分进行序列分析后发现，其存在let-7非依赖的、编码翻译与核糖体装置组分的转录本富集现象，尤其在腺嘌呤/尿嘧啶 (AU) 含量较高的转录本中更为显著；同时神经元发育程序相关转录本则出现明显耗竭。我们进一步证实，LIN28B可通过其冷休克结构域 (cold shock domain, CSD) 和锌指RNA结合结构域 (zinc knuckle domain, ZKD)，优先结合MYCN诱导的核糖体复合物转录本，进而增强其翻译效率。上述数据表明，LIN28B可将MYCN驱动的转录程序与增强的核糖体翻译过程相耦联，从而提示LIN28B是转移表型的转录后调控驱动因子。整体实验设计：对比LIN28B野生型 (wild type, WT) BE2C细胞与LIN28B敲除 (knockout, KO) BE2C细胞，同时在LIN28B KO BE2C细胞中重新表达LIN28B WT、LIN28B突变体（冷休克结构域与锌指结构域完全突变体）以及单突变体LIN28B CSD突变体/ZKD突变体。