The nuclear localization signal of CPSF6 governs post-nuclear import steps of HIV-1 infection

Abstract

The early stages of HIV-1 infection include the trafficking of the viral core into the nucleus of infected cells. However, much remains to be understood about how HIV-1 accomplishes nuclear import and the consequences of the import pathways utilized on nuclear events. The host factor cleavage and polyadenylation specificity factor 6 (CPSF6) assists HIV-1 nuclear localization and post-entry integration targeting. Here, we used a CPSF6 truncation mutant lacking a functional nuclear localization signal (NLS), CPSF6-358, and appended heterologous NLSs to rescue nuclear localization. We show that some, but not all, NLSs drive CPSF6-358 into the nucleus. Interestingly, we found that some nuclear localized CPSF6-NLS chimeras supported inefficient HIV-1 infection. We found that HIV-1 still enters the nucleus in these cell lines but fails to traffic to speckle-associated domains (SPADs). Additionally, we show that HIV-1 fails to efficiently integrate in these cell lines. Collectively, our results demonstrate that the NLS of CPSF6 facilitates steps of HIV-1 infection subsequent to nuclear import and additionally identify the ability of canonical NLS sequences to influence cargo localization in the nucleus following nuclear import. Author SummaryDuring HIV-1 infection, the viral capsid, which encloses the viral genome and accessory proteins required for reverse transcription (RT) and integration, traffics towards the nucleus and enters through the nuclear pore complex (NPC). Following entry into the nucleus, RT is completed and viral capsid disassembles releasing the preintegration complex (PIC) to integrate with the host chromosome. In this study, we investigated the early HIV-1 host factor CPSF6, and specifically focused on the C-terminal short amino acid nuclear localization signal (NLS) in CPSF6, in mediating viral nuclear entry and subsequent gene expression. Altering the NLS in CPSF6 with NLS from other proteins, significantly impacted HIV-1s ability to infect those cells. We further showed this defect in infection occurred at the level of viral integration. This study highlights the importance of the NLS in CPSF6 in dictating the NPC it associates with and its effect on HIV-1 infection. Moreover, our study emphasizes the function of NLS in targeting host cargos to different nuclear entry pathways.