The human CCHC-type Zinc Finger nucleic acid binding protein binds G-rich elements in target mRNA coding sequences initiation promotes translation (RNA-Seq).. Homo sapiens

CNBP is a eukaryote-conserved nucleic-acid binding protein required in mammals for embryonic development. It contains seven CCHC-type zinc-finger domains and was suggested to act as a nucleic acid chaperone, as well as a transcription factor. Here, we identify all CNBP isoforms as cytoplasmic messenger RNA (mRNA)-binding proteins. Using Photoactivatable Ribonucleoside Enhanced Cross-linking and Immunoprecipitation, we mapped its binding sites on RNA at nucleotide-level resolution on a genome-wide scale and find that CNBP interacted with 3961 mRNAs in human cell lines, preferentially at a G-rich motif close to the AUG start codon on mature mRNAs. Loss- and gain-of-function analyses coupled with system-wide RNA and protein quantification revealed that CNBP did not affect RNA abundance, but rather promoted translation of its targets. This is consistent with an RNA chaperone function of CNBP helping to resolve secondary structures, thus promoting translation. Overall design: CNBP protein knockout and RNA-seq

CNBP是一种真核生物保守的核酸结合蛋白，对哺乳动物的胚胎发育不可或缺。该蛋白包含7个CCHC型锌指结构域（CCHC-type zinc-finger domains），此前被推测可同时作为核酸分子伴侣（nucleic acid chaperone）与转录因子（transcription factor）发挥功能。本研究将所有CNBP同工型鉴定为细胞质信使RNA（messenger RNA，mRNA）结合蛋白。借助光激活核糖核苷增强交联免疫沉淀（Photoactivatable Ribonucleoside Enhanced Cross-linking and Immunoprecipitation）技术，我们在全基因组范围内以核苷酸分辨率绘制了其在RNA分子上的结合位点；实验发现，人类细胞系中CNBP可与3961种mRNA结合，且优先结合成熟mRNA上靠近AUG起始密码子的富含G基序。通过功能丧失与功能获得分析，并结合全系统RNA与蛋白质定量实验，我们证实CNBP并不会影响RNA的丰度，而是可促进其靶标mRNA的翻译过程。这一结果与CNBP作为RNA分子伴侣的功能一致：其可帮助解开RNA二级结构，进而促进翻译。整体实验设计：CNBP蛋白敲除与RNA测序（RNA-seq）