The virion-associated incoming HIV-1 RNA genome is not targeted by RNA interference.
BACKGROUND
RNA interference (RNAi) has proven to be a powerful tool to suppress gene expression and can be used as a therapeutic strategy against human pathogenic viruses such as human immunodeficiency virus type 1 (HIV-1). Theoretically, RNAi-mediated inhibition can occur at two points in the replication cycle, upon viral entry before reverse transcription of the RNA genome, and on the newly transcribed viral RNA transcripts. There have been conflicting results on whether RNAi can target the RNA genome of infecting HIV-1 particles. We have addressed this issue with HIV-1-based lentiviral vectors.


RESULTS
We determined the transduction efficiency of a lentiviral vector, as measured by GFP expressing cells, which reflects the number of successful integration events in a cell line stably expressing shNef. We did not observe a difference in the transduction efficiency comparing lentiviral vectors with or without the Nef target sequence in their genome. The results were similar with particles pseudotyped with either the VSV-G or HIV-1 envelope. Additionally, no reduced transduction efficiencies were observed with multiple other shRNAs targeting the vector genome or with synthetic siNef when transiently transfected prior to transduction.


CONCLUSIONS
Our findings indicate that the incoming HIV-1 RNA genome is not targeted by RNAi, probably due to inaccessibility to the RNAi machinery. Thus, therapeutic RNAi strategies aimed at preventing proviral integration should be targeting cellular receptors or co-factors involved in pre-integration events.
Westerhout
Ellen M
EM
Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands. Westerhout: e.m.westerhout@amc.uva.nl
ter Brake
Olivier
O
Berkhout
Ben
B
D003001:Cloning, Molecular; D015693:Gene Products, nef / Q000235:genetics; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics; D016679:Genome, Viral; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics; D015658:HIV Infections / Q000628:therapy; D015497:HIV-1 / Q000235:genetics*; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D012367:RNA, Viral / Q000037:antagonists & inhibitors / Q000235:genetics*; D013601:T-Lymphocytes / Q000502:physiology / Q000821:virology; D014161:Transduction, Genetic; D014771:Virion / Q000235:genetics*; D054311:nef Gene Products, Human Immunodeficiency Virus
D015693:Gene Products, nef; D034741:RNA, Small Interfering; D012367:RNA, Viral; D054311:nef Gene Products, Human Immunodeficiency Virus; D049452:Green Fluorescent Proteins