Dissecting the Cell Entry Pathway of Baculovirus by Single-Particle Tracking and Quantitative Electron Microscopic Analysis

Bombyx mori nucleopolyhedrovirus (BmNPV) is a serious viral pathogen in the sericulture industry and enters host cells via macropinocytic endocytosis; however, the current understanding of the BmNPV entry mechanism remains limited. To confirm whether direct membrane fusion (DMF) results in productive BmNPV infection, DMF infectivity induced by low pH during BmNPV infection was investigated, and the infectious viral particle was traced using an eGFP-labeled virion. We found that BmNPV infection efficiently induced fluid uptake, which allowed BmNPV to bypass the cell membrane barrier via macropinocytosis. However, DMF induced by a low pH abolished the infection. While low pH is an essential condition for membrane fusion triggering, it is not sufficient for productive BmNPV infection, and DMF results in failure to transport the nucleocapsid into the nucleus. These results indicate that transport via macropinocytic vesicles facilitates BmNPV entry into the nucleus and contribute to our understanding of the BmNPV entry mechanism.

Section: Discussionmentioning

confidence: 99%

Transport via Macropinocytic Vesicles Is Crucial for Productive Infection with Bombyx Mori Nucleopolyhedrovirus

Huang

Tang

et al. 2019

“…The pFastBac-Dual donor plasmid (pFD) (Invitrogen, Carlsbad, CA, USA), pMD19-T Simple plasmid (Takara, Tokyo, Japan), pBlueScript plasmid (pBS) (Invitrogen), HST08 competent cells (Takara), DH10B cells containing an AcMNPV bMON14272 bacmid (AcBac) and a pMON7124 helper plasmid (Invitrogen) were used for the experiments. The pFD-VP39-mCherry plasmid was constructed previously in our laboratory [26]. DH10B cells (Invitrogen) containing a vp80 -knockout barmaid were kindly provided by Dr. Zhihong Hu, Wuhan Institute of Virology (WIV), Chinese Academy of Sciences, Wuhan, China.…”

Section: Methodsmentioning

confidence: 99%

A Reverse Genetics System for Cypovirus Based on a Bacmid Expressing T7 RNA Polymerase

Zhang

Yang

Qin

et al. 2019

Self Cite

Dendrolimus punctatus cypovirus (DpCPV), belonging to the genus Cypovirus within the family Reoviridae, is considered the most destructive pest of pine forests worldwide. DpCPV has a genome consisting of 10 linear double-stranded RNA segments. To establish a reverse genetics system, we cloned cDNAs encoding the 10 genomic segments of DpCPV into three reverse genetics vectors in which each segment was transcribed under the control of a T7 RNA polymerase promoter and terminator tagged with a hepatitis delta virus ribozyme sequence. We also constructed a vp80-knockout Autographa californica multiple nucleopolyhedrovirus bacmid to express a T7 RNA polymerase codon-optimized for Sf9 cells. Following transfection of Sf9 cells with the three vectors and the bacmid, occlusion bodies (OBs) with the typical morphology of cypovirus polyhedra were observed by optical microscopy. The rescue system was verified by incorporation of a HindIII restriction enzyme site null mutant of the 9th genomic segment. Furthermore, when we co-transfected Sf9 cells with the reverse genetics vectors, the bacmid, and an additional vector bearing an egfp gene flanked with the 5′ and 3′ untranslated regions of the 10th genomic segment, aggregated green fluorescence co-localizing with the OBs was observed. The rescued OBs were able to infect Spodopetra exigua larvae, although their infectivity was significantly lower than that of wild-type DpCPV. This reverse genetics system for DpCPV could be used to explore viral replication and pathogenesis and to facilitate the development of novel bio-insecticides and expression systems for exogenous proteins.

“…Most if not all animal viruses studied so far manipulate the cellular actin from the first steps of their infection cycle. The binding of virions to specific receptors and entry into host cells is mediated by the cortical network of actin filaments, which are involved in the internalization of the virions localized at the surface of the host cells [ 8 , 9 , 10 ]. The dynamic polymerization of actin reshapes cellular membranes allowing viral internalization and has a role in the formation and movement of endocytic vesicles in case of entry through endocytosis mechanisms [ 11 ].…”

Section: The Cytoskeleton In the Context Of Animal–virus Interactionsmentioning

confidence: 99%

Cross Talk between Viruses and Insect Cells Cytoskeleton

Khorramnejad

Perdomo

Palatini

et al. 2021

Viruses are excellent manipulators of host cellular machinery, behavior, and life cycle, with the host cell cytoskeleton being a primordial viral target. Viruses infecting insects generally enter host cells through clathrin-mediated endocytosis or membrane fusion mechanisms followed by transport of the viral particles to the corresponding replication sites. After viral replication, the viral progeny egresses toward adjacent cells and reaches the different target tissues. Throughout all these steps, actin and tubulin re-arrangements are driven by viruses. The mechanisms used by viruses to manipulate the insect host cytoskeleton are well documented in the case of alphabaculoviruses infecting Lepidoptera hosts and plant viruses infecting Hemiptera vectors, but they are not well studied in case of other insect–virus systems such as arboviruses–mosquito vectors. Here, we summarize the available knowledge on how viruses manipulate the insect host cell cytoskeleton, and we emphasize the primordial role of cytoskeleton components in insect virus motility and the need to expand the study of this interaction.