Dengue Virus NS5 Protein Integrates The Spliceosome And Modulates Cellular Splicing During Infection

DE MAIO, FEDERICO ANDRÉS; RISSO, GUILLERMO; IGLESIAS, NÉSTOR GABRIEL; SHAH, PRIYA; KROGAN, NEVAN; ANDINO, RAÚL; SREBROW, ANABELLA; GAMARNIK, ANDREA V.

Mar del Plata

Congreso; LI Reunión Anual Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2015

Sociedad Argentina de Investigación Bioquímica y Biología Molecular (SAIB)

The plus strand dengue virus RNA genome is translated into a single polyprotein which is then cleaved into 10 different viral proteins. One of these proteins, NS5, contains an RNA-dependent RNA polymerase domain and is essential for viral replication. In order to identify cellular proteins that interact with NS5 during infection, we generated recombinant viruses with a purification tag fused to this viral protein and obtained replication competent viruses. Using an affinity purification and mass spectrometry strategy about 50 cellular proteins were identified as specific NS5 binders. Among these binders, spliceosome components were highly abundant with a clear enrichment of U5 snRNP components. Different studies including co-IP and RIP analysis confirmed the presence of the viral protein in active splicing complexes. Analysis of alternative splicing events from endogenous genes and transfected minigenes indicated that viral infection or NS5 overexpression alters cellular splicing patterns. To explore the relevance of NS5-spliceosome interaction on viral infection, different spliceosomal components were silenced. Interestingly, interfering with U5 components resulted in a significant increase in viral replication. These results support a model in which dengue virus reprograms cellular splicing during infection, creating a more favorable cell environment for viral replication.