TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization

Edward M. Campbell, Jared Weingart, Paola Sette, Silvana Opp, Jaya Sastri, Sarah K. O'Connor, Sarah Talley, Felipe Diaz-Griffero, Vanessa Hirsch, Fadila Bouamrb

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Rhesus macaque TRIM5α (rhTRIM5α) is a retroviral restriction factor that inhibits HIV-1 infection. Previous studies have revealed that TRIM5α restriction occurs via a two-step process. The first step is restriction factor binding, which is sufficient to inhibit infection. The second step, which is sensitive to proteasome inhibition, prevents the accumulation of reverse transcription products in the target cell. However, because of the pleotropic effects of proteasome inhibitors, the molecular mechanisms underlying the individual steps in the restriction process have remained poorly understood. In this study, we have fused the small catalytic domain of herpes simplex virus UL36 deubiquitinase (DUb) to the N-terminal RING domain of rhTRIM5α, which results in a ubiquitination-resistant protein. Cell lines stably expressing this fusion protein inhibited HIV-1 infection to the same degree as a control fusion to a catalytically inactive DUb. However, reverse transcription products were substantially increased in the DUb-TRIM5α fusion relative to the catalytically inactive control or the wild-type (WT) TRIM5α. Similarly, expression of DUb-rhTRIM5α resulted in the accumulation of viral cores in target cells following infection, while the catalytically inactive control and WT rhTRIM5α induced the abortive disassembly of viral cores, indicating a role for ubiquitin conjugation in rhTRIM5α-mediated destabilization of HIV-1 cores. Finally, DUb-rhTRIM5α failed to activate NF-κB signaling pathways compared to controls, demonstrating that this ubiquitination-dependent activity is separable from the ability to restrict retroviral infection.

Original languageEnglish (US)
Pages (from-to)1849-1857
Number of pages9
JournalJournal of Virology
Volume90
Issue number4
DOIs
StatePublished - 2016

Fingerprint

reverse transcription
capsid
Capsid
ubiquitin
Ubiquitin
Human immunodeficiency virus 1
Macaca mulatta
Reverse Transcription
HIV-1
infection
Ubiquitination
proteasome endopeptidase complex
HIV Infections
Infection
herpes simplex
Proteasome Inhibitors
Proteasome Endopeptidase Complex
Simplexvirus
active sites
Catalytic Domain

ASJC Scopus subject areas

  • Immunology
  • Virology

Cite this

Campbell, E. M., Weingart, J., Sette, P., Opp, S., Sastri, J., O'Connor, S. K., ... Bouamrb, F. (2016). TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization. Journal of Virology, 90(4), 1849-1857. https://doi.org/10.1128/JVI.01948-15

TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization. / Campbell, Edward M.; Weingart, Jared; Sette, Paola; Opp, Silvana; Sastri, Jaya; O'Connor, Sarah K.; Talley, Sarah; Diaz-Griffero, Felipe; Hirsch, Vanessa; Bouamrb, Fadila.

In: Journal of Virology, Vol. 90, No. 4, 2016, p. 1849-1857.

Research output: Contribution to journalArticle

Campbell, EM, Weingart, J, Sette, P, Opp, S, Sastri, J, O'Connor, SK, Talley, S, Diaz-Griffero, F, Hirsch, V & Bouamrb, F 2016, 'TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization', Journal of Virology, vol. 90, no. 4, pp. 1849-1857. https://doi.org/10.1128/JVI.01948-15
Campbell, Edward M. ; Weingart, Jared ; Sette, Paola ; Opp, Silvana ; Sastri, Jaya ; O'Connor, Sarah K. ; Talley, Sarah ; Diaz-Griffero, Felipe ; Hirsch, Vanessa ; Bouamrb, Fadila. / TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization. In: Journal of Virology. 2016 ; Vol. 90, No. 4. pp. 1849-1857.
@article{0d529be13a0d428291acdcdbfc5916e5,
title = "TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization",
abstract = "Rhesus macaque TRIM5α (rhTRIM5α) is a retroviral restriction factor that inhibits HIV-1 infection. Previous studies have revealed that TRIM5α restriction occurs via a two-step process. The first step is restriction factor binding, which is sufficient to inhibit infection. The second step, which is sensitive to proteasome inhibition, prevents the accumulation of reverse transcription products in the target cell. However, because of the pleotropic effects of proteasome inhibitors, the molecular mechanisms underlying the individual steps in the restriction process have remained poorly understood. In this study, we have fused the small catalytic domain of herpes simplex virus UL36 deubiquitinase (DUb) to the N-terminal RING domain of rhTRIM5α, which results in a ubiquitination-resistant protein. Cell lines stably expressing this fusion protein inhibited HIV-1 infection to the same degree as a control fusion to a catalytically inactive DUb. However, reverse transcription products were substantially increased in the DUb-TRIM5α fusion relative to the catalytically inactive control or the wild-type (WT) TRIM5α. Similarly, expression of DUb-rhTRIM5α resulted in the accumulation of viral cores in target cells following infection, while the catalytically inactive control and WT rhTRIM5α induced the abortive disassembly of viral cores, indicating a role for ubiquitin conjugation in rhTRIM5α-mediated destabilization of HIV-1 cores. Finally, DUb-rhTRIM5α failed to activate NF-κB signaling pathways compared to controls, demonstrating that this ubiquitination-dependent activity is separable from the ability to restrict retroviral infection.",
author = "Campbell, {Edward M.} and Jared Weingart and Paola Sette and Silvana Opp and Jaya Sastri and O'Connor, {Sarah K.} and Sarah Talley and Felipe Diaz-Griffero and Vanessa Hirsch and Fadila Bouamrb",
year = "2016",
doi = "10.1128/JVI.01948-15",
language = "English (US)",
volume = "90",
pages = "1849--1857",
journal = "Journal of Virology",
issn = "0022-538X",
publisher = "American Society for Microbiology",
number = "4",

}

TY - JOUR

T1 - TRIM5α-mediated ubiquitin chain conjugation is required for inhibition of HIV-1 reverse transcription and capsid destabilization

AU - Campbell, Edward M.

AU - Weingart, Jared

AU - Sette, Paola

AU - Opp, Silvana

AU - Sastri, Jaya

AU - O'Connor, Sarah K.

AU - Talley, Sarah

AU - Diaz-Griffero, Felipe

AU - Hirsch, Vanessa

AU - Bouamrb, Fadila

PY - 2016

Y1 - 2016

N2 - Rhesus macaque TRIM5α (rhTRIM5α) is a retroviral restriction factor that inhibits HIV-1 infection. Previous studies have revealed that TRIM5α restriction occurs via a two-step process. The first step is restriction factor binding, which is sufficient to inhibit infection. The second step, which is sensitive to proteasome inhibition, prevents the accumulation of reverse transcription products in the target cell. However, because of the pleotropic effects of proteasome inhibitors, the molecular mechanisms underlying the individual steps in the restriction process have remained poorly understood. In this study, we have fused the small catalytic domain of herpes simplex virus UL36 deubiquitinase (DUb) to the N-terminal RING domain of rhTRIM5α, which results in a ubiquitination-resistant protein. Cell lines stably expressing this fusion protein inhibited HIV-1 infection to the same degree as a control fusion to a catalytically inactive DUb. However, reverse transcription products were substantially increased in the DUb-TRIM5α fusion relative to the catalytically inactive control or the wild-type (WT) TRIM5α. Similarly, expression of DUb-rhTRIM5α resulted in the accumulation of viral cores in target cells following infection, while the catalytically inactive control and WT rhTRIM5α induced the abortive disassembly of viral cores, indicating a role for ubiquitin conjugation in rhTRIM5α-mediated destabilization of HIV-1 cores. Finally, DUb-rhTRIM5α failed to activate NF-κB signaling pathways compared to controls, demonstrating that this ubiquitination-dependent activity is separable from the ability to restrict retroviral infection.

AB - Rhesus macaque TRIM5α (rhTRIM5α) is a retroviral restriction factor that inhibits HIV-1 infection. Previous studies have revealed that TRIM5α restriction occurs via a two-step process. The first step is restriction factor binding, which is sufficient to inhibit infection. The second step, which is sensitive to proteasome inhibition, prevents the accumulation of reverse transcription products in the target cell. However, because of the pleotropic effects of proteasome inhibitors, the molecular mechanisms underlying the individual steps in the restriction process have remained poorly understood. In this study, we have fused the small catalytic domain of herpes simplex virus UL36 deubiquitinase (DUb) to the N-terminal RING domain of rhTRIM5α, which results in a ubiquitination-resistant protein. Cell lines stably expressing this fusion protein inhibited HIV-1 infection to the same degree as a control fusion to a catalytically inactive DUb. However, reverse transcription products were substantially increased in the DUb-TRIM5α fusion relative to the catalytically inactive control or the wild-type (WT) TRIM5α. Similarly, expression of DUb-rhTRIM5α resulted in the accumulation of viral cores in target cells following infection, while the catalytically inactive control and WT rhTRIM5α induced the abortive disassembly of viral cores, indicating a role for ubiquitin conjugation in rhTRIM5α-mediated destabilization of HIV-1 cores. Finally, DUb-rhTRIM5α failed to activate NF-κB signaling pathways compared to controls, demonstrating that this ubiquitination-dependent activity is separable from the ability to restrict retroviral infection.

UR - http://www.scopus.com/inward/record.url?scp=84958069171&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84958069171&partnerID=8YFLogxK

U2 - 10.1128/JVI.01948-15

DO - 10.1128/JVI.01948-15

M3 - Article

C2 - 26676782

AN - SCOPUS:84958069171

VL - 90

SP - 1849

EP - 1857

JO - Journal of Virology

JF - Journal of Virology

SN - 0022-538X

IS - 4

ER -