FMR1 targets distinct mRNA sequence elements to regulate protein expression [Affymetrix]

Fragile-X Syndrome (FXS) is a multi-organ disease leading to mental retardation, macro-orchidism in males, and premature ovarian insufficiency in female carriers. FXS is also a prominent monogenic disease associated with autism spectrum disorders (ASD). FXS is typically caused by the loss of FRAGILE X-MENTAL RETARDATION 1 (FMR1) expression, which encodes for the RNA-binding protein (RBP), FMR1 (or FMRP). We report the discovery of the RNA recognition elements (RREs), binding sites, and mRNA targets for wild-type and I304N mutant FMRP isoforms as well as its paralogs, FXR1 and FXR2. RRE frequency, ratio, and distribution determine target mRNA association with FMRP. Among highly-enriched targets, we identified many genes involved in ASD and demonstrate that FMRP can affect their protein levels in cell culture, mice, and human brain. Unexpectedly, we discovered that these targets are also dysregulated in Fmr1-/- mouse ovaries, showing signs of premature follicular overdevelopment. These results indicate that FMRP targets shared signaling pathways across different cellular contexts. As it is become increasingly appreciated that signaling pathways are important to FXS and ASD, our results here provide an invaluable molecular guide towards the pursuit of novel therapeutic targets for these devastating neurological disorders. The mRNA profile of RNA recovered from FLAG-antibody immunoprecipitated FMRP was compared to the mRNA profile of the starting lysate material.

脆性X综合征（Fragile-X Syndrome, FXS）是一种多器官疾病，可引发智力障碍、男性巨睾症以及女性携带者的卵巢早衰。脆性X综合征同时也是与自闭症谱系障碍（autism spectrum disorders, ASD）密切相关的典型单基因疾病。该病通常由脆性X智力低下1（FRAGILE X-MENTAL RETARDATION 1, FMR1）基因表达缺失所致，该基因编码RNA结合蛋白（RNA-binding protein, RBP）FMR1（又称FMRP）。本研究报道了野生型及I304N突变型FMRP异构体，以及其旁系同源蛋白FXR1、FXR2的RNA识别元件（RNA recognition elements, RREs）、结合位点与mRNA靶标。RNA识别元件的频率、比例与分布特征决定了靶mRNA与FMRP的结合关联。在高度富集的靶标中，本研究鉴定出众多与自闭症谱系障碍相关的基因，并证实FMRP可在细胞培养体系、小鼠模型及人类大脑中调控这些基因的蛋白表达水平。出乎意料的是，本研究还发现这些靶标在Fmr1基因敲除小鼠的卵巢中同样存在表达失调现象，并呈现出卵泡过早过度发育的特征。上述结果表明，FMRP所靶向的信号通路在不同细胞环境中具有保守性。鉴于信号通路在脆性X综合征与自闭症谱系障碍中的关键作用日益受到重视，本研究结果为探索这两种毁灭性神经疾病的新型治疗靶点提供了极具价值的分子指导。本研究将FLAG抗体免疫沉淀获取的FMRP结合RNA的mRNA表达谱，与初始裂解物的mRNA表达谱进行了对比分析。