A ribonucleoprotein complex protects the interleukin-6 mRNA from degradation by distinct herpesviral endonucleases

PLoS Pathog. 2015 May 12;11(5):e1004899. doi: 10.1371/journal.ppat.1004899. eCollection 2015 May.

Abstract

During lytic Kaposi's sarcoma-associated herpesvirus (KSHV) infection, the viral endonuclease SOX promotes widespread degradation of cytoplasmic messenger RNA (mRNA). However, select mRNAs escape SOX-induced cleavage and remain robustly expressed. Prominent among these is interleukin-6 (IL-6), a growth factor important for survival of KSHV infected B cells. IL-6 escape is notable because it contains a sequence within its 3' untranslated region (UTR) that can confer protection when transferred to a SOX-targeted mRNA, and thus overrides the endonuclease targeting mechanism. Here, we pursued how this protective RNA element functions to maintain mRNA stability. Using affinity purification and mass spectrometry, we identified a set of proteins that associate specifically with the protective element. Although multiple proteins contributed to the escape mechanism, depletion of nucleolin (NCL) most severely impacted protection. NCL was re-localized out of the nucleolus during lytic KSHV infection, and its presence in the cytoplasm was required for protection. After loading onto the IL-6 3' UTR, NCL differentially bound to the translation initiation factor eIF4H. Disrupting this interaction, or depleting eIF4H, reinstated SOX targeting of the RNA, suggesting that interactions between proteins bound to distant regions of the mRNA are important for escape. Finally, we found that the IL-6 3' UTR was also protected against mRNA degradation by the vhs endonuclease encoded by herpes simplex virus, despite the fact that its mechanism of mRNA targeting is distinct from SOX. These findings highlight how a multitude of RNA-protein interactions can impact endonuclease targeting, and identify new features underlying the regulation of the IL-6 mRNA.

Publication types

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

MeSH terms

  • 3' Untranslated Regions
  • B-Lymphocytes / metabolism
  • B-Lymphocytes / pathology
  • B-Lymphocytes / virology
  • Cell Line, Transformed
  • Endonucleases / metabolism*
  • Genes, Reporter
  • HEK293 Cells
  • Half-Life
  • Herpesviridae Infections / metabolism
  • Herpesviridae Infections / pathology
  • Herpesviridae Infections / virology
  • Herpesvirus 8, Human / enzymology*
  • Humans
  • Hydrolysis
  • Interleukin-6 / genetics
  • Interleukin-6 / metabolism*
  • Nucleolin
  • Phosphoproteins / antagonists & inhibitors
  • Phosphoproteins / genetics
  • Phosphoproteins / metabolism*
  • Protein Transport
  • RNA / metabolism
  • RNA Interference
  • RNA Stability*
  • RNA, Messenger / metabolism*
  • RNA-Binding Proteins / antagonists & inhibitors
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Response Elements
  • Ribonucleoproteins / genetics
  • Ribonucleoproteins / metabolism*
  • Viral Proteins / metabolism

Substances

  • 3' Untranslated Regions
  • IL6 protein, human
  • Interleukin-6
  • Phosphoproteins
  • RNA, Messenger
  • RNA, recombinant
  • RNA-Binding Proteins
  • Recombinant Fusion Proteins
  • Ribonucleoproteins
  • Viral Proteins
  • RNA
  • Endonucleases