Identification of cellular cofactors for human immunodeficiency virus replication via a ribozyme-based genomics approach.
Ribozymes are small, catalytic RNA molecules that can be engineered to down-regulate gene expression by cleaving specific mRNA. Here we report the selection of hairpin ribozymes that inhibit human immunodeficiency virus (HIV) replication from a combinatorial ribozyme library. We identified a total of 17 effective ribozymes, each capable of inhibiting HIV infection of human CD4(+) cells. These ribozymes target diverse steps of the viral replication cycle, ranging from entry to transcription. One ribozyme suppressed HIV integration and transcription by inhibiting the expression of the Ku80 subunit of the DNA-activated protein kinase. Another ribozyme specifically inhibited long terminal repeat transactivation, while two additional ones blocked a step that can be bypassed by vesicular stomatitis virus G-protein pseudotyping. The function of Ku80 in HIV replication and its mechanism of action were further confirmed using short interfering RNA. Identification of the gene targets of these and other selected ribozymes may reveal novel therapeutic targets for combating HIV infection.
Waninger
Shani
S
Department of Biological Sciences, Biology Unit 1, Florida State University, Tallahassee, FL 32306-4370, USA.
Kuhen
Kelli
K
Hu
Xiuyuan
X
Chatterton
Jon E
JE
Wong-Staal
Flossie
F
Tang
Hengli
H
D017209:Apoptosis; D015496:CD4-Positive T-Lymphocytes / Q000821:virology; D051747:DNA-Activated Protein Kinase; D004268:DNA-Binding Proteins / Q000502:physiology; D016679:Genome, Viral; D006678:HIV / Q000235:genetics / Q000502:physiology*; D006801:Humans; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D021122:Protein Subunits; D016337:RNA, Catalytic / Q000494:pharmacology*; D034741:RNA, Small Interfering / Q000494:pharmacology; D014779:Virus Replication
D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D021122:Protein Subunits; D016337:RNA, Catalytic; D034741:RNA, Small Interfering; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human; D017346:Protein Serine-Threonine Kinases