We have previously described a C-terminally truncated form of CPSF6 (CPSF6-358) that has potent antiviral activity against different HIV-1, HIV-2, and SIV isolates, but no activity against EIAV, FIV, or MLV. CPSF6-358 interferes with the nuclear entry of HIV-1. We selected for HIV-1 replication in cells expressing CPSF6-358 and identified a mutation of N74D of CA as sufficient to confer viral resistance. We examined the nuclear pore dependencies for wild-type (WT) and N74D HIV-1 and found that the isolates require some of the same pore proteins but differ in requirements for others. In particular, N74D HIV-1 was less sensitive to depletion of TNPO3 (a karyopherin), NUP153, and NUP358/RanBP2 than WT HIV-1. These experiments established a genetic interaction between HIV-1 CA and the nuclear pore. We are now attempting to define the route that HIV-1 takes to the nucleus: the cell factors it interacts with and how it interacts with these factors. Our data suggest a direct interaction of WT CPSF6 with CA. We have also discovered that TNPO3 regulates CPSF6 nuclear localization. We are examining whether CPSF6 normally interacts with HIV-1 CA in the cytoplasm and whether this interaction is important for nuclear entry of the virus. Within the nucleus, we find that the HIV-1 pre-integration complex interacts with CPSF6, and this interaction regulates HIV-1 integration in the host cell chromatin. Notably, we find that by interacting with CPSF6, HIV-1 is first directed to speckles after entering the nucleus and prior to integration. Nuclear speckles colocalize with regions of chromatin that HIV-1 preferentially integrates. Thus, we hypothesize that HIV-1 co-opted CPSF6 to access regions of chromatin that are broadly transcriptionally active due to their proximity to nuclear speckles.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
1ZIABC010487-16
Application #
9779646
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
16
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Basic Sciences
Department
Type
DUNS #
City
State
Country
Zip Code
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