Lentiviral delivery of short hairpin RNAs protects CD4 T cells from multiple clades and primary isolates of HIV.
Viral heterogeneity is a major hurdle for potential therapeutic use of RNA interference (RNAi) against HIV-1. To determine the extent of RNAi tolerance to mutations, we tested 3 viral target sites with differing propensity for mutations: a highly variable rev sequence, a gag sequence conserved only among clade B isolates, and a vif sequence highly conserved across clades. Lentiviral expression of all 3 shRNAs inhibited replication of the homologous HIV(IIIB) strain. However, they differed in their ability to protect primary CD4 T cells against multiple isolates within and across HIV clades. The least conserved rev sequence inhibited only 2 of 5 clade B isolates. The gag sequence (conserved within clade B) protected 5 of 5 clade B isolates but not other clade viruses with 2 or 3 mutations in the central region. In contrast, the vif sequence, which was conserved across clades except for single mutations at positions 14 and 17, inhibited viruses from 5 different clades. Moreover, siRNAs with introduced mutations at sites of gag sequence polymorphisms showed reduced antiviral activity, whereas mutations in vif siRNA only modestly decreased silencing. Thus, although 1 or 2 mutations at peripheral sites are tolerated, mutations in the central target cleavage region abolish RNAi activity.
Lee
Sang-Kyung
SK
The CBR Institute for Biomedical Research, 800 Huntington Ave, Boston, MA 02115, USA.
Dykxhoorn
Derek M
DM
Kumar
Priti
P
Ranjbar
Shahin
S
Song
Erwei
E
Maliszewski
Laura E
LE
François-Bongarçon
Vanessa
V
Goldfeld
Anne
A
Swamy
N Manjunath
NM
Lieberman
Judy
J
Shankar
Premlata
P
D000998:Antiviral Agents / Q000008:administration & dosage / Q000494:pharmacology; D001691:Biological Therapy; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects / Q000821:virology*; D002460:Cell Line; D017124:Conserved Sequence; D015750:Genes, gag; D015757:Genes, rev; D016341:Genes, vif; D015658:HIV Infections / Q000188:drug therapy*; D015497:HIV-1 / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D016086:Lentivirus; D009154:Mutation; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000494:pharmacology; D014162:Transfection
D000998:Antiviral Agents; D034741:RNA, Small Interfering