Pathogen-derived resistance to dengue type 2 virus in mosquito cells by expression of the premembrane coding region of the viral genome.
The full-length premembrane (prM) coding region of the dengue virus type 2 (DEN-2; Jamaica) genome was expressed in C6/36 (Aedes albopictus) cells in either the sense or the antisense orientation from a double subgenomic Sindbis (dsSIN) virus. Northern (RNA) blot analysis confirmed the expression of sense or antisense DEN-2 prM RNA in infected C6/36 cells. PrM protein was demonstrated in cells infected with dsSIN virus expressing DEN-2 sense RNAs by an immunofluorescence assay. C6/36 cells were infected with each dsSIN virus at a multiplicity of infection (MOI) of 50 and challenged 48 h later with DEN-2 virus at an MOI of 0.1. Whereas C6/36 cells infected with a control of dsSIN virus supported high levels of DEN-2 replication, C6/36 cells infected with the dsSIN virus expressing prM antisense RNA were completely resistant to DEN-2 challenge. Cells expressing prM protein or untranslatable prM sense RNA also were resistant to DEN-2 challenge. Cells expressing prM protein demonstrated some breakthrough of DEN-2 virus when challenged at an MOI of 10. However, expressed untranslatable sense prM RNA conferred complete protection to challenge at the high MOI.
Gaines
P J
PJ
Arthropod-Borne and Infectious Diseases Laboratory, Department of Microbiology, Colorado State University, Fort Collins, Colorado 80523, USA.
Olson
K E
KE
Higgs
S
S
Powers
A M
AM
Beaty
B J
BJ
Blair
C D
CD
D000330:Aedes / Q000166:cytology / Q000821:virology; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D006224:Cricetinae; D004279:DNA, Viral; D003716:Dengue Virus / Q000235:genetics* / Q000502:physiology; D015870:Gene Expression; D005818:Genetic Engineering; D016679:Genome, Viral; D006801:Humans; D008969:Molecular Sequence Data; D016372:RNA, Antisense / Q000235:genetics; D012367:RNA, Viral; D012845:Sindbis Virus / Q000235:genetics; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000502:physiology
D004279:DNA, Viral; D016372:RNA, Antisense; D012367:RNA, Viral; D014759:Viral Envelope Proteins; C074932:prM protein, Flavivirus