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Ion-abrasion scanning electron microscopy reveals surface-connected tubular conduits in HIV-infected macrophages
Adam E Bennett1, Kedar Narayan, Dan Shi
1Laboratory of Cell Biology, Center for Cancer Research, NCI, NIH, Bethesda, Maryland, United States of America.
Abstract:
HIV-1-containing internal compartments are readily detected in images of thin sections from infected cells using conventional transmission electron microscopy, but the origin, connectivity, and 3D distribution of these compartments has remained controversial. Here, we report the 3D distribution of viruses in HIV-1-infected primary human macrophages using cryo-electron tomography and ion-abrasion scanning electron microscopy (IA-SEM), a recently developed approach for nanoscale 3D imaging of whole cells. Using IA-SEM, we show the presence of an extensive network of HIV-1-containing tubular compartments in infected macrophages, with diameters of approximately 150-200 nm, and lengths of up to approximately 5 microm that extend to the cell surface from vesicular compartments that contain assembling HIV-1 virions. These types of surface-connected tubular compartments are not observed in T cells infected with the 29/31 KE Gag-matrix mutant where the virus is targeted to multi-vesicular bodies and released into the extracellular medium. IA-SEM imaging also allows visualization of large sheet-like structures that extend outward from the surfaces of macrophages, which may bend and fold back to allow continual creation of viral compartments and virion-lined channels. This potential mechanism for efficient virus trafficking between the cell surface and interior may represent a subversion of pre-existing vesicular machinery for antigen capture, processing, sequestration, and presentation.
Insights
Researchers visualized the 3D distribution of Human Immunodeficiency Virus type 1 (HIV-1) in macrophages. They discovered extensive tubular compartments connecting to the cell surface, facilitating virus trafficking and potentially subverting cellular machinery.
Area of Science:
- Cell Biology
- Virology
- Microscopy
Background:
- Conventional transmission electron microscopy struggles to resolve the 3D structure of HIV-1 internal compartments.
- The origin, connectivity, and distribution of these viral compartments in infected cells remain controversial.
Purpose of the Study:
- To elucidate the 3D distribution and connectivity of Human Immunodeficiency Virus type 1 (HIV-1) containing compartments in primary human macrophages.
- To investigate the role of novel imaging techniques in visualizing nanoscale viral structures within whole cells.
Main Methods:
- Cryo-electron tomography and ion-abrasion scanning electron microscopy (IA-SEM) were employed for nanoscale 3D imaging of whole infected cells.
- IA-SEM enabled visualization of extensive networks of tubular compartments within macrophages.
Main Results:
- An extensive network of HIV-1-containing tubular compartments (150-200 nm diameter, up to 5 µm length) was identified in macrophages, extending from vesicular compartments to the cell surface.
- Surface-connected tubular compartments were absent in T cells infected with a specific HIV-1 Gag-matrix mutant.
- Large, outward-extending sheet-like structures on macrophage surfaces were observed, potentially involved in creating viral compartments and channels.
Conclusions:
- HIV-1 utilizes extensive surface-connected tubular compartments in macrophages for efficient intracellular trafficking.
- This mechanism may involve the subversion of cellular machinery originally involved in antigen processing and presentation.
- IA-SEM is a powerful tool for studying the 3D organization of viruses within host cells.
