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Dengue Virus Impairs Mitochondrial Fusion by Cleaving Mitofusins.

Yu CY, Liang JJ, Li JK, Lee YL, Chang BL, Su CI, Huang WJ, Lai MM, Lin YL - PLoS Pathog. (2015)

Bottom Line: By knockdown and overexpression approaches, these two MFNs showed diverse functions in DENV infection.MFN1 was required for efficient antiviral retinoic acid-inducible gene I-like receptor signaling to suppress DENV replication, while MFN2 participated in maintaining mitochondrial membrane potential (MMP) to attenuate DENV-induced cell death.Thus, MFNs participate in host defense against DENV infection by promoting the antiviral response and cell survival, and DENV regulates mitochondrial morphology by cleaving MFNs to manipulate the outcome of infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Laboratory Science and Biotechnology, National Cheng Kung University, Tainan, Taiwan.

ABSTRACT
Mitochondria are highly dynamic subcellular organelles participating in many signaling pathways such as antiviral innate immunity and cell death cascades. Here we found that mitochondrial fusion was impaired in dengue virus (DENV) infected cells. Two mitofusins (MFN1 and MFN2), which mediate mitochondrial fusion and participate in the proper function of mitochondria, were cleaved by DENV protease NS2B3. By knockdown and overexpression approaches, these two MFNs showed diverse functions in DENV infection. MFN1 was required for efficient antiviral retinoic acid-inducible gene I-like receptor signaling to suppress DENV replication, while MFN2 participated in maintaining mitochondrial membrane potential (MMP) to attenuate DENV-induced cell death. Cleaving MFN1 and MFN2 by DENV protease suppressed mitochondrial fusion and deteriorated DENV-induced cytopathic effects through subverting interferon production and facilitating MMP disruption. Thus, MFNs participate in host defense against DENV infection by promoting the antiviral response and cell survival, and DENV regulates mitochondrial morphology by cleaving MFNs to manipulate the outcome of infection.

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Blocking mitochondrial fusion attenuates RLR signaling and facilitates DENV infection.(A) Mitochondrial morphology analysis of A549 stable cell line overexpressing control GFP or V5-tagged MFN1-T109A. Morphology and subcellular localization of mitochondria and MFN1-T109A were revealed by MitoTracker and anti-V5 staining, respectively. (B) Western blot analysis of GFP- and MFN1-T109A-expressing A549 cells upon DENV infection. Cells were infected with DENV serotype 2 (moi 10) and harvested at indicated time. (C) MMP measurement of vector control (Ctrl) or MFN1-T109A-expressing cells after DENV infection. Cells were infected with DENV (moi 5) for 36 h and harvested for flow cytometry analysis with JC-1 staining. (D to G) Time course study of DENV infection in GFP- and MFN1-T109A-A549 cells. Samples were harvested for the quantification of intracellular IFNβ mRNA (D) and DENV RNA (E) by RT-qPCR, and for the measurement of LDH release (F) and cell viability by trypan blue exclusion (G). Data are mean ± SD (n = 3 per group) and were compared by two-tailed Student’s t test.
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ppat.1005350.g010: Blocking mitochondrial fusion attenuates RLR signaling and facilitates DENV infection.(A) Mitochondrial morphology analysis of A549 stable cell line overexpressing control GFP or V5-tagged MFN1-T109A. Morphology and subcellular localization of mitochondria and MFN1-T109A were revealed by MitoTracker and anti-V5 staining, respectively. (B) Western blot analysis of GFP- and MFN1-T109A-expressing A549 cells upon DENV infection. Cells were infected with DENV serotype 2 (moi 10) and harvested at indicated time. (C) MMP measurement of vector control (Ctrl) or MFN1-T109A-expressing cells after DENV infection. Cells were infected with DENV (moi 5) for 36 h and harvested for flow cytometry analysis with JC-1 staining. (D to G) Time course study of DENV infection in GFP- and MFN1-T109A-A549 cells. Samples were harvested for the quantification of intracellular IFNβ mRNA (D) and DENV RNA (E) by RT-qPCR, and for the measurement of LDH release (F) and cell viability by trypan blue exclusion (G). Data are mean ± SD (n = 3 per group) and were compared by two-tailed Student’s t test.

Mentions: To further address the importance of MFN1 in DENV infection, we established a stable cell line overexpressing a mutated MFN1-T109A [42], which dominant-negatively blocked mitochondrial fusion and resulted in fragmented mitochondria (Fig 10A). Compared to the GFP control cells, DENV-infected MFN1-T109A cells showed lower pIRF3, higher caspase 3 activation (Fig 10B) and more MMP disruption (Fig 10C). Consistently, MFN1-T109A cells induced lower IFNβ mRNA (Fig 10D), and resulted in higher DENV RNA replication (Fig 10E) and severer cytotoxicity measured by LDH release and cell viability (Fig 10F and 10G). Thus, mitochondria fusion contributes to host innate defense against DENV infection.


Dengue Virus Impairs Mitochondrial Fusion by Cleaving Mitofusins.

Yu CY, Liang JJ, Li JK, Lee YL, Chang BL, Su CI, Huang WJ, Lai MM, Lin YL - PLoS Pathog. (2015)

Blocking mitochondrial fusion attenuates RLR signaling and facilitates DENV infection.(A) Mitochondrial morphology analysis of A549 stable cell line overexpressing control GFP or V5-tagged MFN1-T109A. Morphology and subcellular localization of mitochondria and MFN1-T109A were revealed by MitoTracker and anti-V5 staining, respectively. (B) Western blot analysis of GFP- and MFN1-T109A-expressing A549 cells upon DENV infection. Cells were infected with DENV serotype 2 (moi 10) and harvested at indicated time. (C) MMP measurement of vector control (Ctrl) or MFN1-T109A-expressing cells after DENV infection. Cells were infected with DENV (moi 5) for 36 h and harvested for flow cytometry analysis with JC-1 staining. (D to G) Time course study of DENV infection in GFP- and MFN1-T109A-A549 cells. Samples were harvested for the quantification of intracellular IFNβ mRNA (D) and DENV RNA (E) by RT-qPCR, and for the measurement of LDH release (F) and cell viability by trypan blue exclusion (G). Data are mean ± SD (n = 3 per group) and were compared by two-tailed Student’s t test.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4696832&req=5

ppat.1005350.g010: Blocking mitochondrial fusion attenuates RLR signaling and facilitates DENV infection.(A) Mitochondrial morphology analysis of A549 stable cell line overexpressing control GFP or V5-tagged MFN1-T109A. Morphology and subcellular localization of mitochondria and MFN1-T109A were revealed by MitoTracker and anti-V5 staining, respectively. (B) Western blot analysis of GFP- and MFN1-T109A-expressing A549 cells upon DENV infection. Cells were infected with DENV serotype 2 (moi 10) and harvested at indicated time. (C) MMP measurement of vector control (Ctrl) or MFN1-T109A-expressing cells after DENV infection. Cells were infected with DENV (moi 5) for 36 h and harvested for flow cytometry analysis with JC-1 staining. (D to G) Time course study of DENV infection in GFP- and MFN1-T109A-A549 cells. Samples were harvested for the quantification of intracellular IFNβ mRNA (D) and DENV RNA (E) by RT-qPCR, and for the measurement of LDH release (F) and cell viability by trypan blue exclusion (G). Data are mean ± SD (n = 3 per group) and were compared by two-tailed Student’s t test.
Mentions: To further address the importance of MFN1 in DENV infection, we established a stable cell line overexpressing a mutated MFN1-T109A [42], which dominant-negatively blocked mitochondrial fusion and resulted in fragmented mitochondria (Fig 10A). Compared to the GFP control cells, DENV-infected MFN1-T109A cells showed lower pIRF3, higher caspase 3 activation (Fig 10B) and more MMP disruption (Fig 10C). Consistently, MFN1-T109A cells induced lower IFNβ mRNA (Fig 10D), and resulted in higher DENV RNA replication (Fig 10E) and severer cytotoxicity measured by LDH release and cell viability (Fig 10F and 10G). Thus, mitochondria fusion contributes to host innate defense against DENV infection.

Bottom Line: By knockdown and overexpression approaches, these two MFNs showed diverse functions in DENV infection.MFN1 was required for efficient antiviral retinoic acid-inducible gene I-like receptor signaling to suppress DENV replication, while MFN2 participated in maintaining mitochondrial membrane potential (MMP) to attenuate DENV-induced cell death.Thus, MFNs participate in host defense against DENV infection by promoting the antiviral response and cell survival, and DENV regulates mitochondrial morphology by cleaving MFNs to manipulate the outcome of infection.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Laboratory Science and Biotechnology, National Cheng Kung University, Tainan, Taiwan.

ABSTRACT
Mitochondria are highly dynamic subcellular organelles participating in many signaling pathways such as antiviral innate immunity and cell death cascades. Here we found that mitochondrial fusion was impaired in dengue virus (DENV) infected cells. Two mitofusins (MFN1 and MFN2), which mediate mitochondrial fusion and participate in the proper function of mitochondria, were cleaved by DENV protease NS2B3. By knockdown and overexpression approaches, these two MFNs showed diverse functions in DENV infection. MFN1 was required for efficient antiviral retinoic acid-inducible gene I-like receptor signaling to suppress DENV replication, while MFN2 participated in maintaining mitochondrial membrane potential (MMP) to attenuate DENV-induced cell death. Cleaving MFN1 and MFN2 by DENV protease suppressed mitochondrial fusion and deteriorated DENV-induced cytopathic effects through subverting interferon production and facilitating MMP disruption. Thus, MFNs participate in host defense against DENV infection by promoting the antiviral response and cell survival, and DENV regulates mitochondrial morphology by cleaving MFNs to manipulate the outcome of infection.

Show MeSH
Related in: MedlinePlus