Using the massively parallel genetic algorithm for RNA folding, we show that the core region of the 3'-untranslated region of the dengue virus (DENV) RNA can form two dumbbell structures (5'- and 3'-DBs) of unequal frequencies of occurrence. These structures have the propensity to form two potential pseudoknots between identical five-nucleotide terminal loops 1 and 2 (TL1 and TL2) and their complementary pseudoknot motifs, PK2 and PK1. Mutagenesis using a DENV2 replicon RNA encoding the Renilla luciferase reporter indicated that all four motifs and the conserved sequence 2 (CS2) element within the 3'-DB are important for replication. However, for translation, mutation of TL1 alone does not have any effect; TL2 mutation has only a modest effect in translation, but translation is reduced by ∼60% in the TL1/TL2 double mutant, indicating that TL1 exhibits a cooperative synergy with TL2 in translation. Despite the variable contributions of individual TL and PK motifs in translation, WT levels are achieved when the complementarity between TL1/PK2 and TL2/PK1 is maintained even under conditions of inhibition of the translation initiation factor 4E function mediated by LY294002 via a noncanonical pathway. Taken together, our results indicate that the cis-acting RNA elements in the core region of DENV2 RNA that include two DB structures are required not only for RNA replication but also for optimal translation.
|Titolo:||Identification of cis-acting elements in the 3'-untranslated region of the dengue virus type 2 RNA that modulate translation and replication|
|Parole Chiave:||Virus Replication ; Protein Biosynthesis ; Replicon ; Algorithms ; Dengue Virus ; Nucleic Acid Conformation ; RNA, Viral ; Base Sequence ; Conserved Sequence ; Luciferases, Renilla ; Regulatory Sequences, Ribonucleic Acid ; Molecular Sequence Data ; 3' Untranslated Regions ; Cell Line|
|Settore Scientifico Disciplinare:||Settore MED/04 - Patologia Generale|
|Data di pubblicazione:||24-giu-2011|
|Digital Object Identifier (DOI):||10.1074/jbc.M111.234302|
|Appare nelle tipologie:||01 - Articolo su periodico|