The Effect of PF74 on HIV-1 capsid stability and reverse-transcription induced disassembly

The RNA genome of human immunodeficiency virus type 1 (HIV-1) is enclosed
inside a capsid shell that disassembles within a cell in a process known as
uncoating. Using time-lapse atomic force microscopy of purified HIV-1 cores,
we recently showed that the pressure inside the capsid increases during reverse
transcription, which leads to a complete rupture of the capsid near the narrow
end of the cone. High-resolution mechanical mapping revealed the formation of
a stiff coiled filamentous that disappears when the stiffness of the capsid drops.
If the capsid stability is too high (e.g. with the E45A CA mutant), the force
generated during reverse transcription is not sufficient to breach the capsid
structure. We now show that binding of the CA-targeting inhibitor PF74 to
capsids assembled from CA protein in vitro and to HIV-1 isolated cores increases
the stability of the capsid in a concentration-dependent manner. At a PF74
concentration of 10 M, the mechanical stability of the core is elevated to a level
similar to that of the hyperstable capsid mutant E45A. In contrast to HIV-1 cores
without PF74, which undergo complete disassembly after 24 h of reverse
transcription, cores with PF74 only partially disassemble: specifically, the main
body of the capsid remains intact and stiff, but a cap-like structure dissociates
from the narrow end of the core. Moreover, the internal coiled structure is formed
and persists throughout the entire duration of the measurement (~24 h). Our
results provide direct evidence that PF74 directly stabilizes the HIV-1 capsid
lattice, thereby permitting reverse transcription while interfering with a late step
in uncoating.