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Fig. 3 | Retrovirology

Fig. 3

From: Trim28 acts as restriction factor of prototype foamy virus replication by modulating H3K9me3 marks and destabilizing the viral transactivator Tas

Fig. 3

Trim28 negatively regulates Tas-dependent transcriptional activation of PFV LTR and IP promoter activity. a Trim28 inhibited the Tas-dependent transactivation activity of PFV LTR promoter. HEK293T cells seeded in 24-well plates were co-transfected with pCMV-Flag-Trim28 (200 ng or 400 ng, and pCMV-Flag as an empty control), pRL-TK (3 ng), and pGL3-PFV-LTR-luc (70 ng) firefly luciferase reporter. These co-transfected plasmids were combined with or without pTK-Tas (50 ng). b Trim28 inhibited the Tas-dependent transactivation activity of PFV IP promoter. HEK293T cells seeded in 24-well plates were transfected with pCMV-Flag or pCMV-Flag-Trim28 (200 ng or 400 ng), pRL-TK (3 ng), with or without pTK-Tas (20 ng) and pGL3-PFV-IP-luc (20 ng) firefly luciferase reporter. c Trim28 inhibited the Tas-dependent transactivation activity of PFV LTR promoter in a dose dependent way. HEK293T cells seeded in 24-well plates were transfected with pCMV-Flag or pCMV-Flag-Trim28 (0 ~ 800 ng), combined with pRL-TK (3 ng), pTK-Tas (50 ng) and pGL3-PFV-LTR-luc (70 ng) firefly luciferase reporter. d Knockdown of Trim28 with its specific shRNA up-regulates Tas-dependent transactivation of the PFV LTR promoter a dose dependent way. HEK293T cells seeded in 24-well plates were first transfected with pSuper-shNC or pSuper-shTrim28 (0 ~ 800 ng) for 24 h and then cells were cotransfected with pRL-TK (3 ng), pTK-Tas (50 ng) and pPFV-LTR-luc (70 ng) firefly luciferase reporter for 24 h. e Trim28 inhibited the Tas-dependent transactivation activity of PFV IP promoter in a dose dependent way. HEK293T cells seeded in 24-well plates were transfected with pCMV-Flag or pCMV-Flag-Trim28 (0 ~ 800 ng), pRL-TK (3 ng), pTK-Tas (20 ng) and pGL3-PFV-IP-luc (20 ng) firefly luciferase reporter. f Knockdown of Trim28 with its specific shRNA up-regulates Tas-dependent transactivation of the PFV IP promoter a dose dependent way. HEK293T cells seeded in 24-well plates were first transfected with pSuper-shNC or pSuper-shTrim28 (0 ~ 800 ng) for 24 h and then cells were cotransfected with pRL-TK (3 ng), pTK-Tas (20 ng) and pGL3-PFV-IP-luc (20 ng) firefly luciferase reporter. Luciferase activities were measured as described in the Methods. (paired t-test; *p < 0.05, **p < 0.01, ***p < 0.001)

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Retrovirology

ISSN: 1742-4690

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