Molecular Basis for Nucleotide Conservation at the Ends of the Dengue Virus Genome

PLoS Pathogens

Isabelle Varlet¹  Rym Agred¹  Supanee Potisopon¹  Bruno Canard¹  Stéphane Priet¹  Barbara Selisko¹  Yann Thillier²  Corinne Sallamand²  Françoise Debart²  Jean-Jacques Vasseur² 

[1] Aix-Marseille Université, CNRS, AFMB UMR 7257, 163, Marseille, France;IBMM, UMR 5247 CNRS-UM1-UM2, cc 1704 Université Montpellier 2, Montpellier, France

关键词: RNA synthesis;    Polymerases;    Viral genomics;    RNA viruses;    Dengue virus;    Thumbs;    West Nile virus;    Viral structure;   

DOI  :  10.1371/journal.ppat.1002912

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

The dengue virus (DV) is an important human pathogen from the Flavivirus genus, whose genome- and antigenome RNAs start with the strictly conserved sequence pppAG. The RNA-dependent RNA polymerase (RdRp), a product of the NS5 gene, initiates RNA synthesis de novo, i.e., without the use of a pre-existing primer. Very little is known about the mechanism of this de novo initiation and how conservation of the starting adenosine is achieved. The polymerase domain NS5PolDV of NS5, upon initiation on viral RNA templates, synthesizes mainly dinucleotide primers that are then elongated in a processive manner. We show here that NS5PolDV contains a specific priming site for adenosine 5′-triphosphate as the first transcribed nucleotide. Remarkably, in the absence of any RNA template the enzyme is able to selectively synthesize the dinucleotide pppAG when Mn2+ is present as catalytic ion. The T794 to A799 priming loop is essential for initiation and provides at least part of the ATP-specific priming site. The H798 loop residue is of central importance for the ATP-specific initiation step. In addition to ATP selection, NS5PolDV ensures the conservation of the 5′-adenosine by strongly discriminating against viral templates containing an erroneous 3′-end nucleotide in the presence of Mg2+. In the presence of Mn2+, NS5PolDV is remarkably able to generate and elongate the correct pppAG primer on these erroneous templates. This can be regarded as a genomic/antigenomic RNA end repair mechanism. These conservational mechanisms, mediated by the polymerase alone, may extend to other RNA virus families having RdRps initiating RNA synthesis de novo.

【 授权许可】

CC BY