Conway Anne E, Van Nostrand Eric L, Pratt Gabriel A, Aigner Stefan, Wilbert Melissa L, Sundararaman Balaji, Freese Peter, Lambert Nicole J, Sathe Shashank, Liang Tiffany Y, Essex Anthony, Landais Severine, Burge Christopher B, Jones D Leanne, Yeo Gene W
Department of Cellular and Molecular Medicine, University of California at San Diego, La Jolla, CA 92037, USA; Stem Cell Program and Institute for Genomic Medicine, University of California at San Diego, La Jolla, CA 92037, USA; Laboratory of Genetics, Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
Department of Cellular and Molecular Medicine, University of California at San Diego, La Jolla, CA 92037, USA; Stem Cell Program and Institute for Genomic Medicine, University of California at San Diego, La Jolla, CA 92037, USA.
Cell Rep. 2016 Apr 19;15(3):666-679. doi: 10.1016/j.celrep.2016.03.052. Epub 2016 Apr 7.
Human pluripotent stem cells (hPSCs) require precise control of post-transcriptional RNA networks to maintain proliferation and survival. Using enhanced UV crosslinking and immunoprecipitation (eCLIP), we identify RNA targets of the IMP/IGF2BP family of RNA-binding proteins in hPSCs. At the broad region and binding site levels, IMP1 and IMP2 show reproducible binding to a large and overlapping set of 3' UTR-enriched targets. RNA Bind-N-seq applied to recombinant full-length IMP1 and IMP2 reveals CA-rich motifs that are enriched in eCLIP-defined binding sites. We observe that IMP1 loss in hPSCs recapitulates IMP1 phenotypes, including a reduction in cell adhesion and increase in cell death. For cell adhesion, we find IMP1 maintains levels of integrin mRNA specifically regulating RNA stability of ITGB5 in hPSCs. Additionally, we show that IMP1 can be linked to hPSC survival via direct target BCL2. Thus, transcriptome-wide binding profiles identify hPSC targets modulating well-characterized IMP1 roles.
人类多能干细胞（hPSCs）需要对转录后RNA网络进行精确控制，以维持增殖和存活。利用增强型紫外线交联和免疫沉淀技术（eCLIP），我们鉴定了hPSCs中RNA结合蛋白IMP/IGF2BP家族的RNA靶点。在广泛区域和结合位点水平上，IMP1和IMP2显示出与大量重叠的富含3'UTR的靶点有可重复的结合。应用于重组全长IMP1和IMP2的RNA Bind-N-seq揭示了富含CA的基序，这些基序在eCLIP定义的结合位点中富集。我们观察到hPSCs中IMP1的缺失重现了IMP1的表型，包括细胞粘附减少和细胞死亡增加。对于细胞粘附，我们发现IMP1维持整合素mRNA的水平，特异性调节hPSCs中ITGB5的RNA稳定性。此外，我们表明IMP1可通过直接靶点BCL2与hPSC存活相关联。因此，全转录组结合图谱鉴定了调节IMP1特征明确作用的hPSC靶点。
