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

Anne E Conway1, Eric L Van Nostrand2, Gabriel A Pratt3

  • 1Department 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.

Cell Reports
|April 13, 2016
PubMed

Insights

Human pluripotent stem cells rely on RNA-binding proteins like IMP1 for survival and proliferation. This study identifies key RNA targets of IMP1, revealing its role in cell adhesion and survival through specific mRNA regulation.

Area of Science:

  • Molecular Biology
  • Stem Cell Biology
  • RNA Biology

Background:

  • Human pluripotent stem cells (hPSCs) depend on intricate post-transcriptional RNA regulation for maintaining proliferation and survival.
  • The IMP/IGF2BP family of RNA-binding proteins plays a critical role in these regulatory networks within hPSCs.

Purpose of the Study:

  • To identify and characterize the RNA targets of the IMP/IGF2BP family, specifically IMP1 and IMP2, in hPSCs.
  • To elucidate the functional roles of IMP1 in hPSC proliferation, survival, and cell adhesion.

Main Methods:

  • Enhanced UV crosslinking and immunoprecipitation (eCLIP) was employed to map RNA-binding sites of IMP1 and IMP2 in hPSCs.
  • RNA Bind-N-seq was used to determine sequence motifs recognized by recombinant IMP1 and IMP2.
  • Functional assays were conducted following IMP1 knockdown in hPSCs to assess its impact on cell adhesion and survival.

Main Results:

  • eCLIP identified a large, overlapping set of 3' UTR-enriched RNA targets bound by both IMP1 and IMP2.
  • CA-rich motifs were identified as binding preferences for IMP1 and IMP2.
  • Loss of IMP1 in hPSCs led to reduced cell adhesion and increased cell death, associated with altered expression of integrin (ITGB5) and BCL2 mRNAs.

Conclusions:

  • Transcriptome-wide binding profiles reveal specific hPSC targets regulated by IMP1, including ITGB5 and BCL2.
  • IMP1 is crucial for maintaining hPSC adhesion and survival by regulating the stability of specific target mRNAs.
  • These findings highlight the importance of IMP/IGF2BP proteins in hPSC post-transcriptional regulation and cellular homeostasis.

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