Integrative AUF1 PAR-CLIP analysis uncovers AUF1 roles in translation and genome integrity (PAR-CLIP)

We report that AUF1 modulates global mRNA stability and translation, in turn promoting the maintenance of DNA integrity. Overall design: Please see individual series. In short, for AUF1 PAR-CLIP, the four isoforms of AUF1 (p37, p40, p42, and p45) tagged with a Flag epitope were expressed in HEK293 cells. For total RNA-Seq HEK293 cells were transfected with Control siRNA, AUF1 siRNA, Empty Vector, Flag-AUF1 p37, p40, p42, or p45 as well as WI-38 cells were collected at PDL 15 and 55 and also transfected with Control siRNA, AUF1 siRNA, HuR siRNA. For Ribo-Seq HeLa cells were transfected with Control siRNA, AUF1 siRNA, or HuR siRNA.

本研究表明，AUF1可调控全局mRNA稳定性与翻译过程，进而促进DNA完整性的维持。
实验整体设计：详见各数据集系列。
简言之，针对AUF1的光激活核糖核苷交联免疫沉淀测序（PAR-CLIP）实验：将带有Flag表位标记的4种AUF1同工型（p37、p40、p42及p45）在人类胚胎肾293（HEK293）细胞中过表达。
针对总RNA测序（total RNA-Seq）实验：将HEK293细胞分别转染对照小干扰RNA（siRNA）、AUF1 siRNA、空载体以及携带Flag标签的AUF1 p37、p40、p42或p45；同时收集处于第15、55代群体倍增水平（PDL）的WI-38细胞，并对其转染对照siRNA、AUF1 siRNA及HuR siRNA。
针对核糖体测序（Ribo-Seq）实验：将海拉（HeLa）细胞转染对照siRNA、AUF1 siRNA或HuR siRNA。