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The interferon response inhibits HIV particle production by induction of TRIM22.

Barr SD, Smiley JR, Bushman FD - PLoS Pathog. (2008)

Bottom Line: To assess the role of TRIM22 expressed under natural inducing conditions, we compared the effects of interferon in cells depleted for TRIM22 using RNAi and found that HIV particle release was significantly increased in the knockdown, implying that TRIM22 acts as a natural antiviral effector.TRIM22 did not block the release of MLV or EIAV Gag particles.Inhibition was associated with diffuse cytoplasmic staining of HIV Gag rather than accumulation at the plasma membrane, suggesting TRIM22 disrupts proper trafficking.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Microbiology and Immunology, University of Alberta, Alberta Institute for Viral Immunology, Edmonton, Alberta, Canada. stephen.barr@ualberta.ca

ABSTRACT
Treatment of human cells with Type 1 interferons restricts HIV replication. Here we report that the tripartite motif protein TRIM22 is a key mediator. We used transcriptional profiling to identify cellular genes that were induced by interferon treatment and identified TRIM22 as one of the most strongly up-regulated genes. We confirmed, as in previous studies, that TRIM22 over-expression inhibited HIV replication. To assess the role of TRIM22 expressed under natural inducing conditions, we compared the effects of interferon in cells depleted for TRIM22 using RNAi and found that HIV particle release was significantly increased in the knockdown, implying that TRIM22 acts as a natural antiviral effector. Further studies showed that TRIM22 inhibited budding of virus-like particles containing Gag only, indicating that Gag was the target of TRIM22. TRIM22 did not block the release of MLV or EIAV Gag particles. Inhibition was associated with diffuse cytoplasmic staining of HIV Gag rather than accumulation at the plasma membrane, suggesting TRIM22 disrupts proper trafficking. Mutational analyses of TRIM22 showed that the catalytic amino acids Cys15 and Cys18 of the RING domain are required for TRIM22 antiviral activity. These data disclose a pathway by which Type 1 interferons obstruct HIV replication.

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TRIM22 expression blocks release of HIV-1 Gag-only particles.A) Cells were transfected with pGag and pTRIM22 or pcDNA. Cells and Gag-only particles released into the supernatant were isolated 24 hours after transfection and analyzed by Western blotting using an anti-p24CA or anti-actin antibody. B) Cells were pre-treated for 16 hours with 1000 units/ml of IFNβ followed by transfection with pGag. Cells and Gag-only particles released into the supernatant were isolated after 24 hours and analyzed by Western blotting using an anti-p24CA antibody. C) Cells were co-transfected with pR9 or a plasmid expressing MLV GagPol (pMLV) or an EIAV packaging vector (pEIAV) with or without TRIM22 for 24 hours. Cells and Gag-containing particles were isolated and analyzed by Western blotting using an anti-capsid antibody.
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ppat-1000007-g004: TRIM22 expression blocks release of HIV-1 Gag-only particles.A) Cells were transfected with pGag and pTRIM22 or pcDNA. Cells and Gag-only particles released into the supernatant were isolated 24 hours after transfection and analyzed by Western blotting using an anti-p24CA or anti-actin antibody. B) Cells were pre-treated for 16 hours with 1000 units/ml of IFNβ followed by transfection with pGag. Cells and Gag-only particles released into the supernatant were isolated after 24 hours and analyzed by Western blotting using an anti-p24CA antibody. C) Cells were co-transfected with pR9 or a plasmid expressing MLV GagPol (pMLV) or an EIAV packaging vector (pEIAV) with or without TRIM22 for 24 hours. Cells and Gag-containing particles were isolated and analyzed by Western blotting using an anti-capsid antibody.

Mentions: We co-transfected a codon-optimized HIV Gag expression plasmid in the presence or absence of TRIM22 and measured the accumulation of Gag proteins in the cells and in culture supernatants by Western blot (Figure 4A). In the absence of TRIM22, Gag protein was detected in the cell lysate and efficiently released into the supernatant. In the presence of TRIM22, Gag was readily detected in the cell lysate, but not the supernatant. This effect parallels the effect of IFNβ on Gag-only particle release in HOS-CD4/CXCR4 cells (Figure 4B).


The interferon response inhibits HIV particle production by induction of TRIM22.

Barr SD, Smiley JR, Bushman FD - PLoS Pathog. (2008)

TRIM22 expression blocks release of HIV-1 Gag-only particles.A) Cells were transfected with pGag and pTRIM22 or pcDNA. Cells and Gag-only particles released into the supernatant were isolated 24 hours after transfection and analyzed by Western blotting using an anti-p24CA or anti-actin antibody. B) Cells were pre-treated for 16 hours with 1000 units/ml of IFNβ followed by transfection with pGag. Cells and Gag-only particles released into the supernatant were isolated after 24 hours and analyzed by Western blotting using an anti-p24CA antibody. C) Cells were co-transfected with pR9 or a plasmid expressing MLV GagPol (pMLV) or an EIAV packaging vector (pEIAV) with or without TRIM22 for 24 hours. Cells and Gag-containing particles were isolated and analyzed by Western blotting using an anti-capsid antibody.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2279259&req=5

ppat-1000007-g004: TRIM22 expression blocks release of HIV-1 Gag-only particles.A) Cells were transfected with pGag and pTRIM22 or pcDNA. Cells and Gag-only particles released into the supernatant were isolated 24 hours after transfection and analyzed by Western blotting using an anti-p24CA or anti-actin antibody. B) Cells were pre-treated for 16 hours with 1000 units/ml of IFNβ followed by transfection with pGag. Cells and Gag-only particles released into the supernatant were isolated after 24 hours and analyzed by Western blotting using an anti-p24CA antibody. C) Cells were co-transfected with pR9 or a plasmid expressing MLV GagPol (pMLV) or an EIAV packaging vector (pEIAV) with or without TRIM22 for 24 hours. Cells and Gag-containing particles were isolated and analyzed by Western blotting using an anti-capsid antibody.
Mentions: We co-transfected a codon-optimized HIV Gag expression plasmid in the presence or absence of TRIM22 and measured the accumulation of Gag proteins in the cells and in culture supernatants by Western blot (Figure 4A). In the absence of TRIM22, Gag protein was detected in the cell lysate and efficiently released into the supernatant. In the presence of TRIM22, Gag was readily detected in the cell lysate, but not the supernatant. This effect parallels the effect of IFNβ on Gag-only particle release in HOS-CD4/CXCR4 cells (Figure 4B).

Bottom Line: To assess the role of TRIM22 expressed under natural inducing conditions, we compared the effects of interferon in cells depleted for TRIM22 using RNAi and found that HIV particle release was significantly increased in the knockdown, implying that TRIM22 acts as a natural antiviral effector.TRIM22 did not block the release of MLV or EIAV Gag particles.Inhibition was associated with diffuse cytoplasmic staining of HIV Gag rather than accumulation at the plasma membrane, suggesting TRIM22 disrupts proper trafficking.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Microbiology and Immunology, University of Alberta, Alberta Institute for Viral Immunology, Edmonton, Alberta, Canada. stephen.barr@ualberta.ca

ABSTRACT
Treatment of human cells with Type 1 interferons restricts HIV replication. Here we report that the tripartite motif protein TRIM22 is a key mediator. We used transcriptional profiling to identify cellular genes that were induced by interferon treatment and identified TRIM22 as one of the most strongly up-regulated genes. We confirmed, as in previous studies, that TRIM22 over-expression inhibited HIV replication. To assess the role of TRIM22 expressed under natural inducing conditions, we compared the effects of interferon in cells depleted for TRIM22 using RNAi and found that HIV particle release was significantly increased in the knockdown, implying that TRIM22 acts as a natural antiviral effector. Further studies showed that TRIM22 inhibited budding of virus-like particles containing Gag only, indicating that Gag was the target of TRIM22. TRIM22 did not block the release of MLV or EIAV Gag particles. Inhibition was associated with diffuse cytoplasmic staining of HIV Gag rather than accumulation at the plasma membrane, suggesting TRIM22 disrupts proper trafficking. Mutational analyses of TRIM22 showed that the catalytic amino acids Cys15 and Cys18 of the RING domain are required for TRIM22 antiviral activity. These data disclose a pathway by which Type 1 interferons obstruct HIV replication.

Show MeSH