Mutation of the dengue virus type 2 envelope protein heparan sulfate binding sites or the domain III lateral ridge blocks replication in Vero cells prior to membrane fusion

Journal Article · Fri Jul 05 04:00:00 EDT 2013 · Virology

DOI:https://doi.org/10.1016/J.VIROL.2013.03.011· OSTI ID:22436627

Butrapet, Siritorn; Liss, Nathan M. ^[1]; Bennett, Susan L. ^[2]; Luy, Betty E.; Childers, Thomas; Boroughs, Karen L.; Stovall, Janae L.; Calvert, Amanda E. ^[1]; Blair, Carol D. ^[2]; Huang, Claire Y.-H. ^[1]

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1. Division of Vector-Borne Diseases, Centers for Disease Control and Prevention, Fort Collins, CO 80521 (United States)
2. Arthropod-borne and Infectious Diseases Laboratory, Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80523 (United States)

Using an infectious cDNA clone we engineered seven mutations in the putative heparan sulfate- and receptor-binding motifs of the envelope protein of dengue virus serotype 2, strain 16681. Four mutant viruses, KK122/123EE, E202K, G304K, and KKK305/307/310EEE, were recovered following transfection of C6/36 cells. A fifth mutant, KK291/295EE, was recovered from C6/36 cells with a compensatory E295V mutation. All mutants grew in and mediated fusion of virus-infected C6/36 cells, but three of the mutants, KK122/123EE, E202K, G304K, did not grow in Vero cells without further modification. Two Vero cell lethal mutants, KK291/295EV and KKK307/307/310EEE, failed to replicate in DC-SIGN-transformed Raji cells and did not react with monoclonal antibodies known to block DENV attachment to Vero cells. Additionally, both mutants were unable to initiate negative-strand vRNA synthesis in Vero cells by 72 h post-infection, suggesting that the replication block occurred prior to virus-mediated membrane fusion. - Highlights: • Heparan sulfate- and receptor-binding motifs of DENV2 envelope protein were mutated. • Four mutant viruses were isolated—all could fuse C6/36 cells. • Two of these mutants were lethal in Vero cells without further modification. • Lethal mutations were KK291/295EV and KKK305/307/310EEE. • Cell attachment was implicated as the replication block for both mutants.

• OSTI ID: 22436627
• Journal Information: Virology, Journal Name: Virology Journal Issue: 2 Vol. 441; ISSN VIRLAX; ISSN 0042-6822
• Country of Publication: United States
• Language: English

Related Subjects

60 APPLIED LIFE SCIENCES
LETHAL MUTATIONS
MEMBRANES
MONOCLONAL ANTIBODIES
MUTAGENESIS
MUTANTS
RECEPTORS
SULFATES
SYNTHESIS
VIRUSES