Short-lived AUF1 p42-binding mRNAs of RANKL and BCL6 have two distinct instability elements each

PLoS One. 2018 Nov 12;13(11):e0206823. doi: 10.1371/journal.pone.0206823. eCollection 2018.

Abstract

Regulation of mRNA stability by RNA-protein interactions contributes significantly to quantitative aspects of gene expression. We have identified potential mRNA targets of the AU-rich element binding protein AUF1. Myc-tagged AUF1 p42 was induced in mouse NIH/3T3 cells and RNA-protein complexes isolated using anti-myc tag antibody beads. Bound mRNAs were analyzed with Affymetrix microarrays. We have identified 508 potential target mRNAs that were at least 3-fold enriched compared to control cells without myc-AUF1. 22.3% of the enriched mRNAs had an AU-rich cluster in the ARED Organism database, against 16.3% of non-enriched control mRNAs. The enrichment towards AU-rich elements was also visible by AREScore with an average value of 5.2 in the enriched mRNAs versus 4.2 in the control group. Yet, numerous mRNAs were enriched without a high ARE score. The enrichment of tetrameric and pentameric sequences suggests a broad AUF1 p42-binding spectrum at short U-rich sequences flanked by A or G. Still, some enriched mRNAs were highly unstable, as those of TNFSF11 (known as RANKL), KLF10, HES1, CCNT2, SMAD6, and BCL6. We have mapped some of the instability determinants. HES1 mRNA appeared to have a coding region determinant. Detailed analysis of the RANKL and BCL6 3'UTR revealed for both that full instability required two elements, which are conserved in evolution. In RANKL mRNA both elements are AU-rich and separated by 30 bases, while in BCL6 mRNA one is AU-rich and 60 bases from a non AU-rich element that potentially forms a stem-loop structure.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 3' Untranslated Regions / genetics
  • AU Rich Elements / genetics
  • Animals
  • Binding Sites / genetics
  • HEK293 Cells
  • Heterogeneous Nuclear Ribonucleoprotein D0
  • Heterogeneous-Nuclear Ribonucleoprotein D / genetics
  • Heterogeneous-Nuclear Ribonucleoprotein D / metabolism*
  • Humans
  • Mice
  • NIH 3T3 Cells
  • Oligonucleotide Array Sequence Analysis
  • Protein Isoforms / genetics
  • Protein Isoforms / metabolism
  • Proto-Oncogene Proteins c-bcl-6 / genetics*
  • Proto-Oncogene Proteins c-bcl-6 / metabolism
  • RANK Ligand / genetics*
  • RANK Ligand / metabolism
  • RNA Stability / genetics*
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism

Substances

  • 3' Untranslated Regions
  • BCL6 protein, human
  • Bcl6 protein, mouse
  • HNRNPD protein, human
  • Heterogeneous Nuclear Ribonucleoprotein D0
  • Heterogeneous-Nuclear Ribonucleoprotein D
  • Hnrpd protein, mouse
  • Protein Isoforms
  • Proto-Oncogene Proteins c-bcl-6
  • RANK Ligand
  • RNA, Messenger
  • TNFSF11 protein, human
  • Tnfsf11 protein, mouse

Grants and funding

This work was supported by the Swiss National Science Foundation (http://www.snf.ch) with the grant 3100-065435 to LK, and grant 31003A_125193 to PB. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.