Spinoculation Triggers Dynamic Actin and Cofilin Activity That Facilitates HIV-1 Infection of Transformed and Resting CD4 T Cells

Abstract

Centrifugal inoculation, or spinoculation, is widely used in virology research to enhance viral infection. However, the mechanism remained obscure. Using HIV-1 infection of human T cells as a model, we demonstrate that spinoculation triggers dynamic actin and cofilin activity, probably resulting from cellular responses to centrifugal stress. This actin activity also leads to the upregulation of the HIV-1 receptor and coreceptor, CD4 and CXCR4, enhancing viral binding and entry. We also demonstrate that an actin inhibitor, jasplakinolide, diminishes spin-mediated enhancement. In addition, small interfering RNA (siRNA) knockdown of LIMK1, a cofilin kinase, decreases the enhancement. These results suggest that spin-mediated enhancement cannot be explained simply by a virus-concentrating effect; rather, it is coupled with spin-induced cytoskeletal dynamics that promote receptor mobilization, viral entry, and postentry processes. Our results highlight the importance of cofilin and a dynamic cytoskeleton for the initiation of viral infection. Our results also indicate that caution needs to be taken in data interpretation when cells are spinoculated; some of the spin-induced cellular permissiveness may be beyond the natural capacity of an infecting virus.

Spinoculation Enhances HBV Infection in NTCP-Reconstituted Hepatocytes

“…Various mechanisms for this enhancement have been proposed, from ultrastructural changes in the host cell that render it more permissive to viruses [14], to (surprisingly for such low centrifugation speeds) increased deposition of virions on the cell surface [15]. Herein, we established a HepG2-based NTCP-expressing cell line which is permissive to HBV infection, and developed a spinoculation method to enhance HBV infection with the optimized cell culture conditions and viral inoculum size.…”

The Histone Deacetylase Inhibitor Vorinostat (SAHA) Increases the Susceptibility of Uninfected CD4 + T Cells to HIV by Increasing the Kinetics and Efficiency of Postentry Viral Events

“…3A). However, our infection protocol involved a 2-h spinoculation step, which has recently been reported to induce remodeling of the cortical actin barrier and promote viral infection (36). We reasoned that spinoculation might mask the effects of vorinostat on fusion and therefore repeated the experiment in the absence of spinoculation.…”

Distinct Requirements for HIV-Cell Fusion and HIV-mediated Cell-Cell Fusion

“…However, a study by Guo et al (50) has elegantly shown that spinoculation of CD4 ϩ T cells enhances HIV-1 infection by inducing signaling and actin remodeling and that Jasp pretreatment reduces the effect of spinoculation. Although the authors did not examine a direct effect of spinoculation on viral fusion or infection in adherent cells, their results suggest the possibility that diminished FFWO in the presence of actin inhibitors could be related to the spinoculation step employed in our experiments.…”