A role for Nup358/RanBP2 in microtubule-based transport of HIV capsids

About This Grant

PROJECT SUMMARY HIV infection is critically dependent on the transport of the viral genome from the cell periphery to the nucleus. This process is facilitated by the HIV capsid, which serves as a vehicle for the genome and interacts with the host cell’s intracellular transport machinery. However, the mechanisms by which the virus connects to microtubule motors for cytoplasmic transport remain unclear. This research seeks to address these knowledge gaps by investigating the role of the nucleoporin Nup358 (also known as RanBP2), as a potential adaptor for HIV capsid transport. Nup358 localizes to the cytoplasmic face of the nuclear pore complex (NPC), and has been shown to bind the HIV capsid through its cyclophilin-homology domain. Disruption of Nup358 activity inhibits HIV infection, an outcome that has always been interpreted within the context of blocking the nuclear import of HIV. Interestingly, a number of studies have demonstrated that a fraction of Nup358 is also present in the cytoplasm, and is capable of binding both kinesin and dynein motor complexes. Pilot experiments have revealed that Nup358 engages with capsids beyond its cyclophilin-homology domain, recruits kinesin, and facilitates capsid transport with kinesin along microtubules in vitro. These data suggest an additional role for Nup358, serving as a cargo adaptor protein between the capsid and microtubule motors. This hypothesis challenges the current view of Nup358 as solely an NPC component and positions it as a key mediator of early-stage HIV infection. To explore this hypothesis, the project is organized into two specific aims. Aim 1 focuses on elucidating the architecture and transport properties of capsid-Nup358-motor complexes involved in HIV transport. Using interaction assays and in vitro reconstitution, the study will explore how Nup358 stabilizes capsid-motor complexes, activates motor activity, and coordinates motor functions for efficient microtubule-based transport. Aim 2 seeks to validate the physiological roles of capsid-Nup358-motor complexes by disrupting these complexes in cells and testing the effect on infection and subcellular localization of capsids. This research will uncover how the cytoplasmic form of nucleoporin Nup358 facilitates HIV's active transport, and how it links the microtubule network to nuclear targeting, providing insights to guide antiviral development targeting early stages of the HIV life cycle.