h-Prune as a novel binding protein of NS5A that regulates ERK1/2 activation

Hepatitis C virus (HCV) non-structural 5A (NS5A) protein is associated with a wide variety of host signaling pathways by binding to C-terminal polyproline (PxxP) motifs of various proteins. In this study, we used yeast two-hybrid analysis and a GST pull-down assay to screen a novel NS5A interacting...

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Bibliographic Details
Published in:Applied biological chemistry 2016, 59(4), , pp.543-551
Main Authors: Nam, Miyoung, Kim, Cheol-Hee, Kim, Dong-Uk, Lee, Sook-Jeong, Hoe, Kwang-Lae
Format: Article
Language:English
Subjects:
Activation
Alanine
Amino acids
Applied Microbiology
Attenuation
Binding sites
Biological Techniques
Bioorganic Chemistry
C-Terminus
Chemistry
Chemistry and Materials Science
Epidermal growth factor
Exopolyphosphatase
Extracellular signal-regulated kinase
Growth factors
Hepatitis
Hepatitis C
Hepatocytes
Liver
Mutation
Phosphorylation
Polyproline
Proline
Proteins
Signaling
Strong interactions (field theory)
Viruses
Yeasts
농학
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Summary:Hepatitis C virus (HCV) non-structural 5A (NS5A) protein is associated with a wide variety of host signaling pathways by binding to C-terminal polyproline (PxxP) motifs of various proteins. In this study, we used yeast two-hybrid analysis and a GST pull-down assay to screen a novel NS5A interacting protein and elucidate the binding site and cellular signaling by focusing on recombinant human epidermal growth factor (rhEGF)-mediated ERK1/2 activation. Screening a liver cDNA library revealed that h-prune, a member of the DHH (Asp-His-His) protein superfamily, directly interacted with HCV NS5A C-terminus. In particular, a mutation of five proline amino acids to alanine in this region revealed that these two proteins produced strong interaction through this domain. It is known that h-prune possesses a highly conserved DHH motif, which has exopolyphosphatase activity that accelerates hydrolysis of inorganic polyphosphate. A time-chasing analysis after rhEGF treatment demonstrated that h-prune overexpression almost restored NS5A-mediated attenuation of ERK1/2 phosphorylation, but h-prune itself did not alter this signaling. Although the detailed mechanisms need to be clarified, this study demonstrates that h-prune interacts directly with the PxxP motif of the HCV NS5A C-terminus and that this binding alters the rhEGF-mediated ERK1/2 signaling cascade in liver cells.
ISSN:2468-0834
2468-0842
DOI:10.1007/s13765-016-0193-4