After entering a new host cell, the genetic material of the human immunodeficiency virus (HIV) is encased in a multimeric protein shell: the capsid. Encapsulation is thought to facilitate reverse transcription by reducing the available volume for viral enzymes and genetic material, and also to help protect the integrity of the viral genetic material prior to integration in the host genome. However, the capsid is too large to cross the nuclear pore complex, and must disassemble in a process known as uncoating, thus releasing its contents. There is mounting evidence that the timing of uncoating is of critical importance for HIV infectivity, as drugs and capsid protein mutations which affect the capsid`s stability in the cytosol have been found to greatly hinder the infection process.
Reverse transcription, in which a single stranded RNA molecule (ssRNA) is used as a template to synthesize double stranded DNA (dsDNA), is a major step of the retroviral replication cycle which occurs at roughly the same time as HIV uncoating. Since dsDNA is a considerably more rigid molecule than ssDNA, it has been suggested that the resulting confinement forces due to dsDNA pressure on the capsid`s interior could be sufficient and necessary to initiate uncoating.
To explore a possible role for mechanical forces in HIV uncoating, we used Atomic Force Microscopy (AFM) and nanoindentation techniques to study the morphology and mechanical properties of HIV-1 cores in buffered aqueous environment. Our study was focused on several factors known to affect capsid uncoating: capsid protein mutations, a small-molecule inhibitor of HIV-1 infection (PF-74), and the cellular protein Cyclophilin A. In all cases, we found that the capsid`s stiffness was significantly affected, which supports the notion of internal pressure playing a major role in uncoating. This helps shed further light on this important step of retroviral infection.
