Enhanced CLIP Uncovers IMP Protein-RNA Targets in Human Pluripotent Stem Cells Important for Cell Adhesion and Survival

Anne E Conway; Eric L Van Nostrand; Gabriel A Pratt; Stefan Aigner; Melissa L Wilbert; Balaji Sundararaman; Peter Freese; Nicole J Lambert; Shashank Sathe; Tiffany Y Liang; Anthony Essex; Severine Landais; Christopher B Burge; D Leanne Jones; Gene W Yeo

doi:10.1016/j.celrep.2016.03.052

Enhanced CLIP Uncovers IMP Protein-RNA Targets in Human Pluripotent Stem Cells Important for Cell Adhesion and Survival

Cell Rep. 2016 Apr 19;15(3):666-679. doi: 10.1016/j.celrep.2016.03.052. Epub 2016 Apr 7.

Authors

Affiliations

¹ 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.
² 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.
³ 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; Department of Bioinformatics and Systems Biology, University of California at San Diego, La Jolla, CA 92093, USA.
⁴ Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
⁵ Laboratory of Genetics, Salk Institute for Biological Studies, La Jolla, CA 92037, USA.
⁶ Laboratory of Genetics, Salk Institute for Biological Studies, La Jolla, CA 92037, USA; Department of Molecular, Cellular and Developmental Biology, University of California at Los Angeles, Los Angeles, CA 90095, USA; Eli and Edythe Broad Center of Regenerative Medicine and Stem Cell Research, University of California at Los Angeles, Los Angeles, CA 90095, USA. Electronic address: leannejones@ucla.edu.
⁷ 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; Molecular Engineering Laboratory, A(∗)STAR, Singapore 1190777, Singapore; Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 1190777, Singapore. Electronic address: geneyeo@ucsd.edu.

Abstract

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.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

3' Untranslated Regions / genetics
Base Sequence
Cell Adhesion
Cell Survival
Cross-Linking Reagents / metabolism*
Gene Expression Regulation
Human Embryonic Stem Cells / metabolism
Humans
Immunoprecipitation / methods*
Integrins / metabolism
Nucleotide Motifs / genetics
Pluripotent Stem Cells / cytology*
Pluripotent Stem Cells / metabolism*
Protein Binding
Proto-Oncogene Proteins c-bcl-2 / metabolism
RNA / metabolism*
RNA Stability
RNA, Messenger / genetics
RNA, Messenger / metabolism
RNA-Binding Proteins / metabolism

Substances

3' Untranslated Regions
Cross-Linking Reagents
IGF2BP1 protein, human
IGF2BP3 protein, human
Integrins
Proto-Oncogene Proteins c-bcl-2
RNA, Messenger
RNA-Binding Proteins
RNA

Abstract

Publication types

MeSH terms

Substances

Grants and funding