Decoy approach using RNA-DNA chimera oligonucleotides to inhibit the regulatory function of human immunodeficiency virus type 1 Rev protein.
Human immunodeficiency virus type 1 (HIV-1) encodes two regulatory proteins, Tat and Rev, that bind to target RNA sequences. These are the trans-activation responsive (TAR) RNA and the Rev-responsive element (RRE), respectively. The Rev protein shifts RNA synthesis to viral transcripts by binding to the RRE within the env gene. In the present study we prepared a RNA-DNA chimera consisting of 29 or 31 nucleotides to inhibit the Rev regulatory function by means of the decoy approach. The chimera oligonucleotides (anti-Rev oligonucleotides [AROs]) contained an RNA "bubble" structure (13 oligonucleotides; the Rev-binding element in RRE) that bound Rev with a high affinity in an in vitro assay. The controls were RNA-DNA chimera oligonucleotides (negative control oligonucleotides [NCOs]) similar to ARO, but without the bubble structure, that bound with considerably less affinity to Rev. When the inhibitory effects of these decoys on HIV-1 replication were examined, we found that AROs, but no NCOs, reduced more than 90% of the HIV-1 production generated by productively infected human T-cell lines. The production of primary HIV-1 isolates in healthy donor-derived peripheral blood mononuclear cells was also similarly inhibited by AROs. In addition, the induction of viral mRNAs and antigens in latently HIV-1-infected ACH-2 cells by tumor necrosis factor alpha was specifically inhibited by AROs, but not by NCOs. No apparent cytotoxicity was caused by either decoy. Thus, the use of a Rev-binding element-based decoy, the RNA-DNA chimera oligonucleotide, may represent a safer approach to gene therapy for reducing the virus load in HIV-1-infected individuals.
Nakaya
T
T
Section of Serology, Hokkaido University, Sapporo, Japan.
Iwai
S
S
Fujinaga
K
K
Sato
Y
Y
Otsuka
E
E
Ikuta
K
K
D002460:Cell Line; D002470:Cell Survival; D004279:DNA, Viral / Q000235:genetics; D015694:Gene Products, rev / Q000235:genetics* / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D006801:Humans; D009841:Oligonucleotides / Q000235:genetics*; D012333:RNA, Messenger / Q000037:antagonists & inhibitors; D012367:RNA, Viral / Q000235:genetics; D014779:Virus Replication; D054321:rev Gene Products, Human Immunodeficiency Virus
D004279:DNA, Viral; D015694:Gene Products, rev; D009841:Oligonucleotides; D012333:RNA, Messenger; D012367:RNA, Viral; D054321:rev Gene Products, Human Immunodeficiency Virus