Human Cytomegalovirus Tegument Protein UL82 Inhibits STING-Mediated Signaling to Evade Antiviral Immunity
Authors: Pei-Pei Wang
Yu-Zhi Fu
Shan Su
Min-Hua Luo
Wei-Wei Luo
Zhe-Fu Huang
Shu Li
Yan-Yi Wang
Ming-Ming Hu
Yi-Qun Gao
Hong-Bing Shu
Journal: Cell Host & Microbe (2017)
Discipline: MICROBIOLOGY
Keywords: HCMV; MITA; STING; UL82; cGAS
Affiliations: State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, China 430072., Medical Research Institute, Collaborative Innovation Center for Viral Immunology, School of Medicine, Wuhan University, Wuhan, China 430071., Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, China 430071., State Key Laboratory of Virology, College of Life Sciences, Wuhan University, Wuhan, China 430072; Medical Research Institute, Collaborative Innovation Center for Viral Immunology, School of Medicine, Wuhan University, Wuhan, China 430071. Electronic address: .

Abstract:  Recognition of human cytomegalovirus (HCMV) DNA by the cytosolic sensor cGAS initiates STING-dependent innate antiviral responses. HCMV can antagonize host immune responses to promote latency infection. However, it is unknown whether and how HCMV targets the cGAS-STING axis for immune evasion. Here we identified the HCMV tegument protein UL82 as a negative regulator of STING-dependent antiviral responses. UL82 interacted with STING and impaired STING-mediated signaling via two mechanisms. UL82 inhibited the translocation of STING from the ER to perinuclear microsomes by disrupting the STING-iRhom2-TRAPβ translocation complex. UL82 also impaired the recruitment of TBK1 and IRF3 to the STING complex. The levels of downstream antiviral genes induced by UL82-deficient HCMV were higher than those induced by wild-type HCMV. Conversely, wild-type HCMV replicated more efficiently than the UL82-deficient mutant. These findings reveal an important mechanism of immune evasion by HCMV. 

Retracted: No

PubPeer comments:
 The data shown in Fig. B did not correlate with Fig. A, such as 12 h post-infection of HSV, p-TBK1 decreases, but the induction of IFN mRNA did not change much. And in Fig. B, p-TBK1 did not correlate with p-IRF3 and p-p65. The adaptors were phosphorylated without stimulator at 0 h post-infection, are the cells contaminated? ![file] , 
 Thank you for your interest in the study. Fig. B and Fig. A show results of different types of experiments. In Fig. B, phosphorylation of the kinase TBK1 and the transcription factors IRF3 and p65 was detected by immunoblots; In Fig. A, transcription (mRNA levels) of the downstream antiviral genes was measured by qPCR. Phosphorylation of TBK1, IRF3 and p65 is dynamically regulated following viral infection and precedes transcription of the downstream antiviral genes. In addition to TBK1, transcription of the antiviral genes is also regulated by other TBK1-independent pathways. The kinetics of TBK1 phosphorylation and induction of IFN mRNA can be correlated but not necessary temporally consistent. It would be expected that TBK1 is inactivated earlier than the decrease of IFN mRNA level. The results would be more clear if more time points are done. TBK1, IRF3 and p65 phosphorylation is also temporally and dynamically regulated. TBK1 auto-phosphorylation occurs immediately after its recruitment to MITA STING, IRF3 is phosphorylated by TBK1 thereafter during a trafficking process of the MITA-associated complex, and p65 is phosphorylated by IKK. In addition, the stability of these phosphorylated signaling components is differentially regulated. It is not unexpected that the levels of these phosphorylated components do not correlate at all time points. As you said, the signaling components were phosphorylated to low levels in un-infected cells. This was not caused by cell contamination. It is most likely due to the low basal activity in these RNAi-transduced stable cell lines and or low affinities of the phosphor-specific antibodies to the corresponding un-phosphorylated proteins. Please let me know if you have additional questions. Hong-Bing Shu (shuh@whu.edu.cn) , 
 Letu2019s focus on 96h. KD of STING only enhance viral replication about 2 times, this canu2019t be true, as STING is very critical for IFN-I production, and HCMV is DNA virus. As they claimed that UL82 is very important for viral replication, how come KO of UL82 only decreased viral replication about 3 times. When we do plague assay, 2-3 times difference between different wells is very often, so the data is not convincing. ![file] , 
 #3 HCMV evolves many ways to antagonize the immune system, which is responsible for its persistent infection in even STING MITA-positive and IFN-inducible cells. Therefore, it is not surprising that knockdown of STING MITA enhanced HCMV replication for a couple of times. In addition, the knockdown efficiency of STING MITA and the possible basal IFN activity in these RNAi-transduced cell lines may also contribute to the results. UL82 plays a role in replication does not necessary mean it is required for replication. The data tell how important it is. Hong-Bing Shu , 
 The phosphorylation of p65 shown in Fig. 2B did not correlate with sFig. 3D. The p-p65 level would upregulate 6hrs after HCMV infection (red box). But in the Con group shown in sFig.3D (green box), the peak of p-p65 delayed to 12hrs. I will be very grateful if authors could provide a reasonable explanation. Also, I noticed UL82-siRNA#1 also showed blocking effect on inducible UL44 expression (blue box), could authors explain why? ![file] , 
 #5 These experiments were performed in different cell lines, which respond to HCMV infection with different kinetics. HFF cells were used in Fig. 2B. THP1 cells were used in Fig. S3D. UL82-siRNA#1 did not inhibit UL44 expression. The corresponding b-actin loading control levels were a little bit lower. Yu-Zhi Fu and Hong-Bing Shu ,