Department of Genetics, Washington University School of Medicine, St. Louis, MO 63110, USA; Department of Psychiatry, Washington University School of Medicine, St. Louis, MO 63110, USA.
Department of Genetics, Washington University School of Medicine, St. Louis, MO 63110, USA.
Cell Rep. 2020 Dec 22;33(12):108531. doi: 10.1016/j.celrep.2020.108531.
CELF6 is a CELF-RNA-binding protein, and thus part of a protein family with roles in human disease; however, its mRNA targets in the brain are largely unknown. Using cross-linking immunoprecipitation and sequencing (CLIP-seq), we define its CNS targets, which are enriched for 3' UTRs in synaptic protein-coding genes. Using a massively parallel reporter assay framework, we test the consequence of CELF6 expression on target sequences, with and without mutating putative binding motifs. Where CELF6 exerts an effect on sequences, it is largely to decrease RNA abundance, which is reversed by mutating UGU-rich motifs. This is also the case for CELF3-5, with a protein-dependent effect on magnitude. Finally, we demonstrate that targets are derepressed in CELF6-mutant mice, and at least two key CNS proteins, FOS and FGF13, show altered protein expression levels and localization. Our works find, in addition to previously identified roles in splicing, that CELF6 is associated with repression of its CNS targets via the 3' UTR in vivo.
CELF6 是 CELF-RNA 结合蛋白，因此是在人类疾病中起作用的蛋白家族的一部分；然而，其在大脑中的 mRNA 靶标在很大程度上是未知的。我们使用交联免疫沉淀和测序（CLIP-seq）来定义其 CNS 靶标，这些靶标富含突触蛋白编码基因的 3'UTR。我们使用大规模平行报告基因检测框架来测试 CELF6 表达对靶序列的影响，同时突变可能的结合基序。在 CELF6 对序列产生影响的情况下，它主要是降低 RNA 丰度，而突变 UGU 丰富基序则可以逆转这种情况。CELF3-5 也是如此，其对幅度的影响具有蛋白依赖性。最后，我们证明在 CELF6 突变小鼠中靶标被解除抑制，至少两种关键的中枢神经系统蛋白，FOS 和 FGF13，显示出改变的蛋白表达水平和定位。除了先前在剪接中鉴定的作用外，我们的工作还发现 CELF6 通过体内 3'UTR 与 CNS 靶标的抑制有关。
