Ebola virus VP35 phosphorylation and its role in the viral lifecycle

Tuesday, September 23, 2014 — Poster Session III
12:00 p.m. – 2:00 p.m.	FAES Academic Center	NIAID	VIROL-5

Authors

• MR Kumar
• L Diaz
• S Altmann
• B Ork
• J Kindrachuk
• RF Johnson
• PB Jahrling

Abstract

The Ebola virus (EBOV) protein VP35 is one of seven structural proteins encoded by the viral genome, with roles in viral transcription, replication, genome packaging and the modulation of host interferon responses. The mechanisms by which VP35 is regulated are poorly understood for these multiple processes. In this study, we investigated the role of VP35’s phosphorylation status in regulating EBOV transcription. Following site-directed mutagenesis of VP35 at T15A, S174A, Y176A and S266A, we compared the reporter activity and protein expression of VP35 alanine mutants to wild type VP35 in the GFP-based Ebola minigenome (EMG) reporter assay. We found that several constructs with various mutant combinations demonstrated reduced GFP expression levels. Protein expression of these constructs was unaffected by the mutations, indicating that the change in activation was due to changes in protein activity, not availability. To investigate if charge conservation impacted VP35 function we generated VP35 aspartate mutants at the same residues. The data indicate a phosphorylation pattern which modulates expression of GFP from the EMG assay, suggesting that phosphorylation of specific residues regulates VP35 function. The requirement for post-translation modification of VP35 for regulation of its transcriptional activity indicates that VP35 phosphorylation is a prime target for antiviral therapy.

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