HIV-1 capsid uncoating initiates after the first strand transfer of reverse transcription

Abstract

BackgroundCorrect disassembly of the HIV-1 capsid shell, called uncoating, is increasingly recognised as central for multiple steps during retroviral replication. However, the timing, localisation and mechanism of uncoating are poorly understood and progress in this area is hampered by difficulties in measuring the process. Previous work suggested that uncoating occurs soon after entry of the viral core into the cell, but recent studies report later uncoating, at or in the nucleus. Furthermore, inhibiting reverse transcription delays uncoating, linking these processes. ResultsHere, we have used a combined approach of experimental interrogation of viral mutants and mathematical modelling to investigate the timing of uncoating with respect to reverse transcription. By developing a minimal, testable, model and employing multiple uncoating assays to overcome the disadvantages of each single assay, we find that uncoating is not concomitant with the initiation of reverse transcription. Instead, uncoating appears to be triggered once reverse transcription reaches a certain stage, namely shortly after first strand transfer.ConclusionsUsing multiple approaches, we have identified a point during reverse transcription that induces uncoating of the HIV-1 CA shell. We propose that uncoating initiates after the first strand transfer of reverse transcription.Electronic supplementary materialThe online version of this article (doi:10.1186/s12977-016-0292-7) contains supplementary material, which is available to authorized users.

Reverse Transcription Mechanically Initiates HIV-1 Capsid Disassembly

“…An early report suggested that reverse transcription is required for HIV-1 uncoating in target cells (13). Subsequently, Hulme et al reported that inhibition of reverse transcription can delay uncoating (14); this conclusion was supported by subsequent workers using complementary cellbased assays (15,16). In a more recent study employing a novel fluorescence-based HIV-1 imaging assay, uncoating was observed to accelerate upon the initiation of reverse transcription (17).…”

Early cytoplasmic uncoating is associated with infectivity of HIV-1

“…2B). In recent years, several groups have discovered that it is possible to modulate "uncoating" of HIV-1 particles either by delaying or blocking reverse transcription, or by introducing certain mutations into the CA protein that forms the conical core (2,23,28,29,53,54). A well-established mechanism to disrupt HIV capsid stability is to exploit the mechanism of TRIM5 restriction that recruits the proteasome to direct capsid disruption (16,44).…”

“…An emerging hypothesis to the link between early reverse transcription products and the loss of capsid integrity has been recently backed by a study based on both theoretical and mutational studies. Cosnefroy et al (29) suggested that the first-strand transfer of reverse transcription might play a crucial role in core disassembly. However, another concurrent report that measured the binding of tetrameric CypA to the capsid as a tool to follow CA pointed to no correlation between first-strand transfer and uncoating (27).…”

“…Infection in MDMs might be expected to result in a slower capsid integrity loss since it has been reported that macrophages have relatively low dNTP concentrations (40,48,49) and delayed completion of reverse transcription (28). However, the imaging assay has suggested that capsid integrity loss is associated with early reverse transcription, which might be less sensitive to dNTP concentrations (29,50). Consistent with this interpretation, we show that the timing of capsid integrity loss in macrophages can be modulated by transitory NVP treatment (Fig.…”

“…A third proposed model is that uncoating takes place in the cytoplasm as the reverse-transcribing viral genome is transported to the nucleus. This model is based on both fluorescence microscopy observations and assays that exploit the impact on infection from modulated TRIM5 susceptibility (25)(26)(27)(28)(29). The HIV-1 capsid uncoating dependence on microtubule stability also supports a cytoplasmic uncoating (25,26,(30)(31)(32)(33)(34).…”