Quantitative microscopy of functional HIV post-entry complexes reveals association of replication with the viral capsid
- Heidelberg University, Germany
- German Center for Infection Research, Germany
- Gilead Sciences Inc., United States
- University Heidelberg, Germany
Abstract
The steps from HIV-1 cytoplasmic entry until integration of the reverse transcribed genome are currently enigmatic. They occur in ill-defined reverse-transcription- and pre-integration-complexes (RTC, PIC) with various host and viral proteins implicated. Here, we report quantitative detection of functional RTC/PIC by labeling nascent DNA combined with detection of viral integrase. We show that the viral capsid (CA) protein remains associated with cytoplasmic RTC/PIC, but is lost on nuclear PIC in a HeLa-derived cell line. In contrast, nuclear PIC were almost always CA-positive in primary human macrophages, indicating nuclear import of capsids or capsid-like structures. We further show that the CA-targeted inhibitor PF74 exhibits a bimodal mechanism, blocking RTC/PIC association with the host factor CPSF6 and nuclear entry at low, and abrogating reverse transcription at high concentrations. The newly developed system is ideally suited for studying retroviral post-entry events and the roles of host factors including DNA sensors and signaling molecules.
Article and author information
Author details
Reviewing Editor
- Wesley I Sundquist, University of Utah, United States
Version history
- Received: July 22, 2014
- Accepted: December 15, 2014
- Accepted Manuscript published: December 17, 2014 (version 1)
- Version of Record published: January 15, 2015 (version 2)
Copyright
© 2014, Peng et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Quantitative microscopy of functional HIV post-entry complexes reveals association of replication with the viral capsid
eLife 3:e04114.
https://doi.org/10.7554/eLife.04114
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