TY - JOUR TI - Quantitative microscopy of functional HIV post-entry complexes reveals association of replication with the viral capsid AU - Peng, Ke AU - Muranyi, Walter AU - Glass, Bärbel AU - Laketa, Vibor AU - Yant, Stephen R AU - Tsai, Luong AU - Cihlar, Tomas AU - Müller, Barbara AU - Kräusslich, Hans-Georg A2 - Sundquist, Wesley I VL - 3 PY - 2014 DA - 2014/12/17 SP - e04114 C1 - eLife 2014;3:e04114 DO - 10.7554/eLife.04114 UR - https://doi.org/10.7554/eLife.04114 AB - 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. In this study, we report quantitative detection of functional RTC/PIC by labeling nascent DNA combined with detection of viral integrase. We show that the viral CA (capsid) 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. KW - HIV KW - viral entry KW - reverse transcription KW - macrophage KW - CPSF6 JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -