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The human antiviral factor TRIM11 is under the regulation of HIV-1 Vpr.

Yuan T, Yao W, Huang F, Sun B, Yang R - PLoS ONE (2014)

Bottom Line: In this study, we demonstrated that TRIM11 mainly suppressed the early steps of HIV-1 transduction, resulting in decreased reverse transcripts.Additionally, we found that TRIM11 could inhibit HIV-1 long terminal repeat (LTR) activity, which may be related to its inhibitory effects on NF-κB.These results suggest that the antiviral factor TRIM11 is indirectly regulated by HIV-1 Vpr through unknown mechanisms and that the concentration of Vpr is essential to these processes.

View Article: PubMed Central - PubMed

Affiliation: Research Group of HIV Molecular Epidemiology and Virology, Center for Emerging Infectious Diseases, The State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei, The People's Republic of China.

ABSTRACT
TRIM11 has been reported to be able to restrict HIV-1 replication, but the detailed aspects of the interfering mechanisms remain unclear. In this study, we demonstrated that TRIM11 mainly suppressed the early steps of HIV-1 transduction, resulting in decreased reverse transcripts. Additionally, we found that TRIM11 could inhibit HIV-1 long terminal repeat (LTR) activity, which may be related to its inhibitory effects on NF-κB. Deletion mutant experiments showed that the RING domain of TRIM11 was indispensable in inhibiting the early steps of HIV-1 transduction but was dispensable in decreasing NF-κB and LTR activities. Moreover, we found that low levels of Vpr decreased TRIM11 protein levels, while high levels increased them, and these regulations were independent of the VprBP-associated proteasome machinery. These results suggest that the antiviral factor TRIM11 is indirectly regulated by HIV-1 Vpr through unknown mechanisms and that the concentration of Vpr is essential to these processes. Thus, our work confirms TRIM11 as a host cellular factor that interferes with the early steps of HIV-1 replication and provides a connection between viral protein and host antiviral factors.

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Effects of the RING domain and proteasome system on the functioning of TRIM11.Lysates from HEK293 cells stably transduced with pCDH, pCDH-TRIM11 or pCDH-RDTRIM11 were subjected to a western blot with the indicated antibodies. B. HEK293 cells stably expressing TRIM11, RDTRIM11 or a control pCDH vector were inoculated with various amounts of HIV-1 Vpr− viruses. Luciferase assays were performed at 24 hpi. C. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses and were analyzed by qPCR for viral DNA. D. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were pretreated with MG132 or DMSO for 5 h and inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses. Luciferase assays were performed at 14 hpi. E, F. HEK293 cells were cotransfected with 900 ng of plasmids expressing TRIM11, RDTRIM11 or control vector along with 50 ng of the HIV-1 LTR firefly luciferase reporter (E) or 50 ng of the NF-κB firefly luciferase reporter (F) and 50 ng of renilla luciferase plasmid. Luciferase assays were performed at 24 h after transfection. Error bars represent the standard deviations from three independent replicates of the same experiment.
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pone-0104269-g004: Effects of the RING domain and proteasome system on the functioning of TRIM11.Lysates from HEK293 cells stably transduced with pCDH, pCDH-TRIM11 or pCDH-RDTRIM11 were subjected to a western blot with the indicated antibodies. B. HEK293 cells stably expressing TRIM11, RDTRIM11 or a control pCDH vector were inoculated with various amounts of HIV-1 Vpr− viruses. Luciferase assays were performed at 24 hpi. C. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses and were analyzed by qPCR for viral DNA. D. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were pretreated with MG132 or DMSO for 5 h and inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses. Luciferase assays were performed at 14 hpi. E, F. HEK293 cells were cotransfected with 900 ng of plasmids expressing TRIM11, RDTRIM11 or control vector along with 50 ng of the HIV-1 LTR firefly luciferase reporter (E) or 50 ng of the NF-κB firefly luciferase reporter (F) and 50 ng of renilla luciferase plasmid. Luciferase assays were performed at 24 h after transfection. Error bars represent the standard deviations from three independent replicates of the same experiment.

Mentions: TNFα is known be able to activate HIV-1 transcription [11], [36]. Therefore, HEK293 cells were stimulated with TNFα for an additional 4 h after transfection with the TRIM11 overexpression vector or control vector and with the HIV-1 LTR luciferase reporter construct for 24 h. Increasing amounts of TRIM11 also led to decreases in TNFα-stimulated HIV-1 LTR activity (Figure 3C). To determine whether the inhibitory effects of TRIM11 on TNFα-induced LTR activation could be due to impaired basal transcription (Figure 3A), we examined the fold increase in luciferase activity that was observed upon stimulation in both the control and TRIM11- transfected cells. In comparison with the control cells, the TRIM11-overexpressing cells had significantly lower LTR activities in terms of fold induction versus baseline (Figure 4D). These results indicate that TRIM11 inhibited both basal and TNFα-induced HIV-1 LTR transcription.


The human antiviral factor TRIM11 is under the regulation of HIV-1 Vpr.

Yuan T, Yao W, Huang F, Sun B, Yang R - PLoS ONE (2014)

Effects of the RING domain and proteasome system on the functioning of TRIM11.Lysates from HEK293 cells stably transduced with pCDH, pCDH-TRIM11 or pCDH-RDTRIM11 were subjected to a western blot with the indicated antibodies. B. HEK293 cells stably expressing TRIM11, RDTRIM11 or a control pCDH vector were inoculated with various amounts of HIV-1 Vpr− viruses. Luciferase assays were performed at 24 hpi. C. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses and were analyzed by qPCR for viral DNA. D. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were pretreated with MG132 or DMSO for 5 h and inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses. Luciferase assays were performed at 14 hpi. E, F. HEK293 cells were cotransfected with 900 ng of plasmids expressing TRIM11, RDTRIM11 or control vector along with 50 ng of the HIV-1 LTR firefly luciferase reporter (E) or 50 ng of the NF-κB firefly luciferase reporter (F) and 50 ng of renilla luciferase plasmid. Luciferase assays were performed at 24 h after transfection. Error bars represent the standard deviations from three independent replicates of the same experiment.
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Related In: Results  -  Collection

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pone-0104269-g004: Effects of the RING domain and proteasome system on the functioning of TRIM11.Lysates from HEK293 cells stably transduced with pCDH, pCDH-TRIM11 or pCDH-RDTRIM11 were subjected to a western blot with the indicated antibodies. B. HEK293 cells stably expressing TRIM11, RDTRIM11 or a control pCDH vector were inoculated with various amounts of HIV-1 Vpr− viruses. Luciferase assays were performed at 24 hpi. C. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses and were analyzed by qPCR for viral DNA. D. HEK293 cells stably expressing TRIM11, RDTRIM11 or control pCDH vector were pretreated with MG132 or DMSO for 5 h and inoculated with 50 ng/ml (p24gag) of HIV-1 Vpr− viruses. Luciferase assays were performed at 14 hpi. E, F. HEK293 cells were cotransfected with 900 ng of plasmids expressing TRIM11, RDTRIM11 or control vector along with 50 ng of the HIV-1 LTR firefly luciferase reporter (E) or 50 ng of the NF-κB firefly luciferase reporter (F) and 50 ng of renilla luciferase plasmid. Luciferase assays were performed at 24 h after transfection. Error bars represent the standard deviations from three independent replicates of the same experiment.
Mentions: TNFα is known be able to activate HIV-1 transcription [11], [36]. Therefore, HEK293 cells were stimulated with TNFα for an additional 4 h after transfection with the TRIM11 overexpression vector or control vector and with the HIV-1 LTR luciferase reporter construct for 24 h. Increasing amounts of TRIM11 also led to decreases in TNFα-stimulated HIV-1 LTR activity (Figure 3C). To determine whether the inhibitory effects of TRIM11 on TNFα-induced LTR activation could be due to impaired basal transcription (Figure 3A), we examined the fold increase in luciferase activity that was observed upon stimulation in both the control and TRIM11- transfected cells. In comparison with the control cells, the TRIM11-overexpressing cells had significantly lower LTR activities in terms of fold induction versus baseline (Figure 4D). These results indicate that TRIM11 inhibited both basal and TNFα-induced HIV-1 LTR transcription.

Bottom Line: In this study, we demonstrated that TRIM11 mainly suppressed the early steps of HIV-1 transduction, resulting in decreased reverse transcripts.Additionally, we found that TRIM11 could inhibit HIV-1 long terminal repeat (LTR) activity, which may be related to its inhibitory effects on NF-κB.These results suggest that the antiviral factor TRIM11 is indirectly regulated by HIV-1 Vpr through unknown mechanisms and that the concentration of Vpr is essential to these processes.

View Article: PubMed Central - PubMed

Affiliation: Research Group of HIV Molecular Epidemiology and Virology, Center for Emerging Infectious Diseases, The State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, Hubei, The People's Republic of China.

ABSTRACT
TRIM11 has been reported to be able to restrict HIV-1 replication, but the detailed aspects of the interfering mechanisms remain unclear. In this study, we demonstrated that TRIM11 mainly suppressed the early steps of HIV-1 transduction, resulting in decreased reverse transcripts. Additionally, we found that TRIM11 could inhibit HIV-1 long terminal repeat (LTR) activity, which may be related to its inhibitory effects on NF-κB. Deletion mutant experiments showed that the RING domain of TRIM11 was indispensable in inhibiting the early steps of HIV-1 transduction but was dispensable in decreasing NF-κB and LTR activities. Moreover, we found that low levels of Vpr decreased TRIM11 protein levels, while high levels increased them, and these regulations were independent of the VprBP-associated proteasome machinery. These results suggest that the antiviral factor TRIM11 is indirectly regulated by HIV-1 Vpr through unknown mechanisms and that the concentration of Vpr is essential to these processes. Thus, our work confirms TRIM11 as a host cellular factor that interferes with the early steps of HIV-1 replication and provides a connection between viral protein and host antiviral factors.

Show MeSH
Related in: MedlinePlus