Entry of Tiger Frog Virus (an Iridovirus) into HepG2 Cells via a pH-Dependent, Atypical, Caveola-Mediated Endocytosis Pathway

Guo, Chang-Jun; Liu, Dong; Wu, Yanling; Yang, Xiaobo; Yang, Lishi; Mi, Shu; Huang, Yuxin; Luo, Yongwen; Jia, Kuntong; Liu, Zhaoyu; Chen, Wei-Jian; Weng, Shaoping; Yu, Xiao‐Qiang; He, Jianguo

doi:10.1128/jvi.01500-10

Cited by 48 publications

(64 citation statements)

References 67 publications

(96 reference statements)

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“…Dynamin is a 100-kDa GTPase that plays an essential role in cellular membrane fission during vesicle formation and therefore is required for clathrinand caveola-mediated endocytosis (33,(39)(40)(41)(42). Dynamin has been shown to be required for the internalization of numerous viruses (43)(44)(45)(46).…”

Section: Resultsmentioning

confidence: 99%

Entry of Classical Swine Fever Virus into PK-15 Cells via a pH-, Dynamin-, and Cholesterol-Dependent, Clathrin-Mediated Endocytic Pathway That Requires Rab5 and Rab7

Shi

Liu

Zhou

et al. 2016

J Virol

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Classical swine fever virus (CSFV), a member of the genus Pestivirus within the family Flaviviridae, is a small, enveloped, positive-strand RNA virus. Due to its economic importance to the pig industry, the biology and pathogenesis of CSFV have been investigated extensively. However, the mechanisms of CSFV entry into cells are not well characterized. In this study, we used systematic approaches to dissect CSFV cell entry. We first observed that CSFV infection was inhibited by chloroquine and NH 4 Cl, suggesting that viral entry required a low-pH environment. By using the specific inhibitor dynasore, or by expressing the dominant negative (DN) K44A mutant, we verified that dynamin is required for CSFV entry. CSFV particles were observed to colocalize with clathrin at 5 min postinternalization, and CSFV infection was significantly reduced by chlorpromazine treatment, overexpression of a dominant negative form of the EPS15 protein, or knockdown of the clathrin heavy chain by RNA interference. These results suggested that CSFV entry depends on clathrin. Additionally, we found that endocytosis of CSFV was dependent on membrane cholesterol, while neither the overexpression of a dominant negative caveolin mutant nor the knockdown of caveolin had an effect. These results further suggested that CSFV entry required cholesterol and not caveolae. Importantly, the effect of DN mutants of three Rab proteins that regulate endosomal traffic on CSFV infection was examined. Expression of DN Rab5 and Rab7 mutants, but not the DN Rab11 mutant, significantly inhibited CSFV replication. These results were confirmed by silencing of Rab5 and Rab7. Confocal microscopy showed that virus particles colocalized with Rab5 or Rab7 during the early phase of infection within 45 min after virus entry. These results indicated that after internalization, CSFV moved to early and late endosomes before releasing its RNA. Taken together, our findings demonstrate for the first time that CSFV enters cells through the endocytic pathway, providing new insights into the life cycle of pestiviruses. IMPORTANCEBovine viral diarrhea virus (BVDV), a single-stranded, positive-sense pestivirus within the family Flaviviridae, is internalized by clathrin-dependent receptor-mediated endocytosis. However, the detailed mechanism of cell entry is unknown for other pestiviruses, such as classical swine fever (CSF) virus (CSFV). CSFV is the etiological agent of CSF, a highly contagious disease of swine that causes numerous deaths in pigs and enormous economic losses in China. Understanding the entry pathway of CSFV will not only advance our knowledge of CSFV infection and pathogenesis but also provide novel drug targets for antiviral intervention. Based on this objective, we used systematic approaches to dissect the pathway of entry of CSFV into PK-15 cells. This is the first report to show that the entry of CSFV into PK-15 cells requires a low-pH environment and involves dynamin-and cholesteroldependent, clathrin-mediated endocytosis that requires Rab5 and Rab7....

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Section: Resultsmentioning

confidence: 99%

Entry of Classical Swine Fever Virus into PK-15 Cells via a pH-, Dynamin-, and Cholesterol-Dependent, Clathrin-Mediated Endocytic Pathway That Requires Rab5 and Rab7

Shi

Liu

Zhou

et al. 2016

J Virol

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“…For example, cholesterol depletion by treatment with MbCD was shown to result in a 100-fold suppression of West Nile virus infection, whereas addition of exogenous cholesterol restored the infection (49), similar to the finding in this study. Cholesterol is important for generating and maintaining the structure of caveolae, the critical endocytic vesicle used by many viruses, such as SV40 and tiger frog virus, to enter host cells via endocytosis (50,51). By using MbCD to block infection and showing the colocalization of virions with early endosomes, a recent study claimed that WSSV also adopts caveolae-mediated endocytosis to break through the membrane barrier (8).…”

Section: Discussionmentioning

confidence: 99%

Collaboration between a Soluble C-Type Lectin and Calreticulin Facilitates White Spot Syndrome Virus Infection in Shrimp

Wang

et al. 2014

The Journal of Immunology

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White spot syndrome virus (WSSV) mainly infects crustaceans through the digestive tract. Whether C-type lectins (CLs), which are important receptors for many viruses, participate in WSSV infection in the shrimp stomach remains unknown. In this study, we orally infected kuruma shrimp Marsupenaeus japonicus to model the natural transmission of WSSV and identified a CL (designated as M. japonicus stomach virus–associated CL [MjsvCL]) that was significantly induced by virus infection in the stomach. Knockdown of MjsvCL expression by RNA interference suppressed the virus replication, whereas exogenous MjsvCL enhanced it. Further analysis by GST pull-down and coimmunoprecipitation showed that MjsvCL could bind to viral protein 28, the most abundant and functionally relevant envelope protein of WSSV. Furthermore, cell-surface calreticulin was identified as a receptor of MjsvCL, and the interaction between these proteins was a determinant for the viral infection–promoting activity of MjsvCL. The MjsvCL–calreticulin pathway facilitated virus entry likely in a cholesterol-dependent manner. This study provides insights into a mechanism by which soluble CLs capture and present virions to the cell-surface receptor to facilitate viral infection.

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“…56,65 The variance between SHEP and HepG2 cells metabolism, growth rate and cell morphologies cause these inhibitors to display different expression profiles. The pre-treatment of genistein, a tyrosine kinase inhibitor that disrupts the actin network of and cells and inhibits dynamin II the master regulator of membrane trafficking, 66 demonstrated contrasting uptake profiles for both cell lines. The rearrangement of the actin cytoskeleton and disruption of ligand triggered uptake in SHEP cells decreased the cellular internalization profile of P(MAA-co-CMA)-DOX nanocomplexes by 54.4%.…”

Section: The Endocytic Profile Of P(maa-co-cma)-dox Nanocomplexesmentioning

confidence: 99%

The endocytic pathway and therapeutic efficiency of doxorubicin conjugated cholesterol-derived polymers

et al. 2015

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Previously synthesized poly(methacrylic acid-co-cholesteryl methacrylate) P(MAA-co-CMA) copolymers were examined as potential drug delivery vehicles. P(MAA-co-CMA) copolymers were fluorescently labelled and imaged in SHEP and HepG2 cells. To understand their cell internalization pathway endocytic inhibition studies were conducted. It was concluded that P(MAA-co-CMA) are taken up by the cells via clathrin-independent endocytosis (CIE) (both caveolae mediated and cholesterol dependent endocytosis) mechanisms. The formation and characterization of P(MAA-co-CMA)-doxorubicin (DOX) nanocomplexes was investigated by fluorescence lifetime imaging microscopy (FLIM), UV-Visible spectroscopy (UV-Vis) and dynamic light scattering (DLS) studies. The toxicity screening between P(MAA-co-CMA)-DOX nanocomplexes (at varying w/w ratios) and free DOX, revealed nanocomplexes to exhibit higher cytotoxicity towards cancer cells in comparison to normal cells. FLIM and confocal microscopy were employed for investigating the time-dependent release of DOX in SHEP cells and the cellular uptake profile of P(MAAco-CMA)-DOX nanocomplexes in cancer and normal cell lines, respectively. The endocytic pathway of P(MAA-co-CMA)-DOX nanocomplexes were examined in SHEP and HepG2 cells via flow cytometry revealing the complexes to be internalized through both clathrin-dependent (CDE) and CIE mechanisms. The drug delivery profile, reported herein, illuminates the specific endocytic route and therapeutic efficiency of P(MAA-co-CMA)-DOX nanocomplexes strongly suggesting these particles to be promising candidates for in vivo applications.

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Entry of Tiger Frog Virus (an Iridovirus) into HepG2 Cells via a pH-Dependent, Atypical, Caveola-Mediated Endocytosis Pathway

Cited by 48 publications

References 67 publications

Entry of Classical Swine Fever Virus into PK-15 Cells via a pH-, Dynamin-, and Cholesterol-Dependent, Clathrin-Mediated Endocytic Pathway That Requires Rab5 and Rab7

Entry of Classical Swine Fever Virus into PK-15 Cells via a pH-, Dynamin-, and Cholesterol-Dependent, Clathrin-Mediated Endocytic Pathway That Requires Rab5 and Rab7

Collaboration between a Soluble C-Type Lectin and Calreticulin Facilitates White Spot Syndrome Virus Infection in Shrimp

The endocytic pathway and therapeutic efficiency of doxorubicin conjugated cholesterol-derived polymers

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