Reduction of gene expression by a hairpin-loop structured oligodeoxynucleotide: alternative to siRNA and antisense.
BACKGROUND
We previously described the inhibition of HIV-1 replication by a 54-mer hairpin-loop structured oligodeoxynucleotide (ODN) A, which binds the polypurine tract (PPT) on HIV-1 RNA. ODN A was shown to lead to reduced viral RNA in virions or early during infection.


RESULTS
Here we demonstrated that ODN A was able to cause hydrolysis of viral RNA not only by retroviral RT-associated RNase H but also cellular RNase H1 and RNase H2 in vitro. Furthermore, ODN A reduced gene expression in a dose-dependent manner in a cell-based reporter assay where a PPT sequence was inserted in the 5' untranslated region of the reporter gene. The efficacy of ODN A was higher than that of its siRNA and antisense counterparts. By knocking down cellular RNases H, we showed that RNase H1 contributed to the gene silencing by ODN A but the possibility of a partial contribution of RNase H-independent mechanisms could not be ruled out.


CONCLUSIONS
Our findings highlight the potential application of hairpin-loop structured ODNs for reduction of gene expression in mammalian cells and underscore the possibility of using ODN A to trigger the hydrolysis of HIV RNA in infected cells by cellular RNases H.
Kwok
Terry
T
Institute of Medical Virology, University of Zurich, Gloriastrasse 30, CH 8006 Zurich, Switzerland.
Heinrich
Jochen
J
Jung-Shiu
Jiunshan
J
Meier
Michelle G
MG
Mathur
Srikanth
S
Moelling
Karin
K
D001483:Base Sequence; D002460:Cell Line; D002461:Cell Line, Transformed; D005434:Flow Cytometry; D015967:Gene Expression Regulation, Viral; D020868:Gene Silencing; D054303:HIV Reverse Transcriptase / Q000235:genetics / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics / Q000502:physiology; D006801:Humans; D006868:Hydrolysis; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016376:Oligonucleotides, Antisense / Q000235:genetics*; D011072:Poly U / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016914:Ribonuclease H / Q000235:genetics / Q000378:metabolism
D009838:Oligodeoxyribonucleotides; D016376:Oligonucleotides, Antisense; D034741:RNA, Small Interfering; D012367:RNA, Viral; D011072:Poly U; D054303:HIV Reverse Transcriptase; D016914:Ribonuclease H; C078856:ribonuclease HI