Specific interaction between eEF1A and HIV RT is critical for HIV-1 reverse transcription and a potential anti-HIV target

Li, Dongsheng, Wei, Ting, Rawle, Daniel J., Qin, Fangyun, Wang, Rui, Soares, Dinesh C., Jin, Hongping, Sivakumaran, Haran, Lin, Min-Hsuan, Spann, Kirsten, Abbott, Catherine M. and Harrich, David (2015) Specific interaction between eEF1A and HIV RT is critical for HIV-1 reverse transcription and a potential anti-HIV target. PLoS Pathogens, 11 12: e1005289.1-e1005289.12. doi:10.1371/journal.ppat.1005289


Author Li, Dongsheng
Wei, Ting
Rawle, Daniel J.
Qin, Fangyun
Wang, Rui
Soares, Dinesh C.
Jin, Hongping
Sivakumaran, Haran
Lin, Min-Hsuan
Spann, Kirsten
Abbott, Catherine M.
Harrich, David
Title Specific interaction between eEF1A and HIV RT is critical for HIV-1 reverse transcription and a potential anti-HIV target
Journal name PLoS Pathogens   Check publisher's open access policy
ISSN 1553-7366
Publication date 2015-12-01
Sub-type Article (original research)
DOI 10.1371/journal.ppat.1005289
Open Access Status DOI
Volume 11
Issue 12
Start page e1005289.1
End page e1005289.12
Total pages 27
Place of publication San Francisco, CA, United States
Publisher Public Library of Science
Collection year 2016
Language eng
Abstract Reverse transcription is the central defining feature of HIV-1 replication. We previously reported that the cellular eukaryotic elongation factor 1 (eEF1) complex associates with the HIV-1 reverse transcription complex (RTC) and the association is important for late steps of reverse transcription. Here we show that associationbetween the eEF1 and RTC complexes occurs by a strong and direct interaction between the subunit eEF1A and reverse transcriptase (RT). Using biolayer interferometry and co-immunoprecipitation (co-IP) assays, we show that association between the eEF1 and RTC complexes occurs by a strong (KD ~3–4 nM) and direct interaction between eEF1A and reverse transcriptase (RT). Biolayer interferometry analysis of cell lysates with titrated levels of eEF1A indicates it is a predominant cellular RT binding protein. Both the RT thumb and connection domains are required for interaction with eEF1A. A single amino acid mutation, W252A, within the thumb domain impaired co-IP between eEF1A and RT, and also significantly reduced the efficiency of late reverse transcription and virus replication when incorporated into infectious HIV-1. Molecular modeling analysis indicated that interaction between W252 and L303 are important for RT structure, and their mutation to alanine did not impair heterodimerisation, but negatively impacted interaction with eEF1A. Didemnin B, which specifically binds eEF1A, potently inhibited HIV-1 reverse transcription by greater than 2 logs at subnanomolar concentrations, especially affecting reverse transcription late DNA synthesis. Analysis showed reduced levels of RTCs from HIV-1-infected HEK293T treated with didemnin B compared to untreated cells. Interestingly, HIV-1 with a W252A RT mutation was resistant to didemnin B negative effects showing that didemnin B affects HIV-1 by targeting the RT-eEF1A interaction. The combined evidence indicates a direct interaction between eEF1A and RT is crucial for HIV reverse transcription and replication, and the RT-eEF1A interaction is a potential drug target.
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2016 Collection
School of Chemistry and Molecular Biosciences
Child Health Research Centre Publications
 
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Created: Fri, 04 Dec 2015, 10:38:03 EST by Mrs Louise Nimwegen on behalf of School of Chemistry & Molecular Biosciences