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Published January 2014 | Published + Supplemental Material
Journal Article Open

Electron Tomography of HIV-1 Infection in Gut-Associated Lymphoid Tissue


Critical aspects of HIV-1 infection occur in mucosal tissues, particularly in the gut, which contains large numbers of HIV-1 target cells that are depleted early in infection. We used electron tomography (ET) to image HIV-1 in gut-associated lymphoid tissue (GALT) of HIV-1–infected humanized mice, the first three-dimensional ultrastructural examination of HIV-1 infection in vivo. Human immune cells were successfully engrafted in the mice, and following infection with HIV-1, human T cells were reduced in GALT. Virions were found by ET at all stages of egress, including budding immature virions and free mature and immature viruses. Immuno-electron microscopy verified the virions were HIV-1 and showed CD4 sequestration in the endoplasmic reticulum of infected cells. Observation of HIV-1 in infected GALT tissue revealed that most HIV-1–infected cells, identified by immunolabeling and/or the presence of budding virions, were localized to intestinal crypts with pools of free virions concentrated in spaces between cells. Fewer infected cells were found in mucosal regions and the lamina propria. The preservation quality of reconstructed tissue volumes allowed details of budding virions, including structures interpreted as host-encoded scission machinery, to be resolved. Although HIV-1 virions released from infected cultured cells have been described as exclusively mature, we found pools of both immature and mature free virions within infected tissue. The pools could be classified as containing either mostly mature or mostly immature particles, and analyses of their proximities to the cell of origin supported a model of semi-synchronous waves of virion release. In addition to HIV-1 transmission by pools of free virus, we found evidence of transmission via virological synapses. Three-dimensional EM imaging of an active infection within tissue revealed important differences between cultured cell and tissue infection models and furthered the ultrastructural understanding of HIV-1 transmission within lymphoid tissue.

Additional Information

© 2014 Ladinsky et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Received August 11, 2013; Accepted December 9, 2013; Published January 30, 2014. We thank Wes Sundquist for providing anti-ESCRT and other antibodies; Yunji Wu and the Caltech Protein Expression Center for purified 2G12; Drs. Grant Jensen and Julia Greer for use of the Tecnai T12 and F30 electron microscopes, respectively, and Dr. Alasdair McDowall and Carol Garland for help maintaining the microscopes. We thank Ariane Briegel, Wes Sundquist, Bruce Walker, Cora Woodward and Mark Yeager for helpful discussions, and Bruce Walker, Grant Jensen and Cora Woodward for critical reading of the manuscript. Funding: This work was supported by the National Institutes of Health (2 P50 GM082545-06; WI Sundquist, PI), a Ragon Institute post-doctoral fellowship (CK), and gifts from the Gordon and Betty Moore Foundation and the Agouron Institute to support electron microscopy at Caltech. DSK is supported by an NIH K08 Award (K08 AI084546-04) and a fellowship from the Burroughs Wellcome Fund. The MGH Humanized Mouse Program is supported by the Harvard University Center for AIDS Research (P30 AI060354). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Author Contributions: Conceived and designed the experiments: MSL DSK PJB. Performed the experiments: MSL GO. Analyzed the data: MSL CK GO DSK PJB. Contributed reagents/materials/analysis tools: GO MD VV AMT DSK. Wrote the paper: MSL CK DSK PJB.

Attached Files

Published - journal.ppat.1003899.pdf

Supplemental Material - journal.ppat.1003899.s001.tif

Supplemental Material - journal.ppat.1003899.s002.tif

Supplemental Material - journal.ppat.1003899.s003.tif

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Supplemental Material - journal.ppat.1003899.s009.mov

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Supplemental Material - journal.ppat.1003899.s011.mov

Supplemental Material - journal.ppat.1003899.s012.mov

Supplemental Material - journal.ppat.1003899.s013.mov

Supplemental Material - journal.ppat.1003899.s014.docx


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