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The C-terminal amyloidogenic peptide contributes to self-assembly of Avibirnavirus viral protease.

Zheng X, Jia L, Hu B, Sun Y, Zhang Y, Gao X, Deng T, Bao S, Xu L, Zhou J - Sci Rep (2015)

Bottom Line: Interestingly, the last C-terminal hydrophobic and amyloidogenic stretch (238)YHLAMA(243) with two "aggregation-prone" alanine residues was found to be essential for its intracellular self-assembly.Importantly, the assembly of VP4 significantly reduced the cytotoxicity of protease activity in host cells which potentially prevent the premature cell death and facilitate viral replication.This study provides novel insights into the formation mechanism and biological functions of the Avibirnavirus protease-related fibrils.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou 310058, PR China.

ABSTRACT
Unlike other viral protease, Avibirnavirus infectious bursal disease virus (IBDV)-encoded viral protease VP4 forms unusual intracellular tubule-like structures during viral infection. However, the formation mechanism and potential biological functions of intracellular VP4 tubules remain largely elusive. Here, we show that VP4 can assemble into tubules in diverse IBDV-infected cells. Dynamic analysis show that VP4 initiates the assembly at early stage of IBDV infection, and gradually assembles into larger size of fibrils within the cytoplasm and nucleus. Intracellular assembly of VP4 doesn't involve the host cytoskeleton, other IBDV-encoded viral proteins or vital subcellular organelles. Interestingly, the last C-terminal hydrophobic and amyloidogenic stretch (238)YHLAMA(243) with two "aggregation-prone" alanine residues was found to be essential for its intracellular self-assembly. The assembled VP4 fibrils show significantly low solubility, subsequently, the deposition of highly assembled VP4 structures ultimately deformed the host cytoskeleton and nucleus, which was potentially associated with IBDV lytic infection. Importantly, the assembly of VP4 significantly reduced the cytotoxicity of protease activity in host cells which potentially prevent the premature cell death and facilitate viral replication. This study provides novel insights into the formation mechanism and biological functions of the Avibirnavirus protease-related fibrils.

No MeSH data available.


Related in: MedlinePlus

Expression and subcellular dynamics of VP4 in IBDV-infected cells.(a) Subcellular expression of VP4 in IBDV-infected CEF (24 h), DF-1 cells (24 h), Vero cells (72 h), 293T cells (24 h) and intrabursal lymphocytes (72 h). (b) Assembly dynamics of VP4 in IBDV-infected DF-1 cells was examined at 4, 6, 8 and 10 h after infection (MOI = 1) by IFA using anti-VP4 mAb, and nuclei were stained with DAPI. (c) Expression dynamics of VP4 in IBDV-infected DF-1 cells (MOI = 1) was examined at 0, 2, 4, 6, 8 and 10 h after infection by Western blotting using anti-VP4 mAb.
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f1: Expression and subcellular dynamics of VP4 in IBDV-infected cells.(a) Subcellular expression of VP4 in IBDV-infected CEF (24 h), DF-1 cells (24 h), Vero cells (72 h), 293T cells (24 h) and intrabursal lymphocytes (72 h). (b) Assembly dynamics of VP4 in IBDV-infected DF-1 cells was examined at 4, 6, 8 and 10 h after infection (MOI = 1) by IFA using anti-VP4 mAb, and nuclei were stained with DAPI. (c) Expression dynamics of VP4 in IBDV-infected DF-1 cells (MOI = 1) was examined at 0, 2, 4, 6, 8 and 10 h after infection by Western blotting using anti-VP4 mAb.

Mentions: VP4 was previously reported to form tubule-like structures within IBDV-infected chicken embryo fibroblast cells (CEFs)17, but it’s unclear if the intracellular formation of tubular VP4 is specific for certain type of cells. We characterized VP4 expression in various IBDV-infected cells, including primary CEFs, DF-1 cells, Vero cells, HEK293T cells (in vitro) and intrabursal lymphocytes in chicken (in vivo) by immunofluorescence assay (IFA) using anti-VP4 monoclonal antibody (mAb). Confocal microcopy revealed that VP4 assembles within the cytoplasm and nucleus of various IBDV-infected cells (Fig. 1a). Specifically, intranuclear VP4 assembled into varying sizes of fibrils among IBDV-infected different cells, cytoplasmic VP4 in IBDV-infected CEF, DF-1 cells and HEK293T cells mainly formed needle-like fibrils, while cytoplasmic VP4 in the IBDV-infected Vero cells and intrabursal lymphoid cells formed large mass-like aggregates. These data demonstrated that intracellular VP4 assembly is an intrinsic feature of IBDV infection which is independent of cell types.


The C-terminal amyloidogenic peptide contributes to self-assembly of Avibirnavirus viral protease.

Zheng X, Jia L, Hu B, Sun Y, Zhang Y, Gao X, Deng T, Bao S, Xu L, Zhou J - Sci Rep (2015)

Expression and subcellular dynamics of VP4 in IBDV-infected cells.(a) Subcellular expression of VP4 in IBDV-infected CEF (24 h), DF-1 cells (24 h), Vero cells (72 h), 293T cells (24 h) and intrabursal lymphocytes (72 h). (b) Assembly dynamics of VP4 in IBDV-infected DF-1 cells was examined at 4, 6, 8 and 10 h after infection (MOI = 1) by IFA using anti-VP4 mAb, and nuclei were stained with DAPI. (c) Expression dynamics of VP4 in IBDV-infected DF-1 cells (MOI = 1) was examined at 0, 2, 4, 6, 8 and 10 h after infection by Western blotting using anti-VP4 mAb.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4594098&req=5

f1: Expression and subcellular dynamics of VP4 in IBDV-infected cells.(a) Subcellular expression of VP4 in IBDV-infected CEF (24 h), DF-1 cells (24 h), Vero cells (72 h), 293T cells (24 h) and intrabursal lymphocytes (72 h). (b) Assembly dynamics of VP4 in IBDV-infected DF-1 cells was examined at 4, 6, 8 and 10 h after infection (MOI = 1) by IFA using anti-VP4 mAb, and nuclei were stained with DAPI. (c) Expression dynamics of VP4 in IBDV-infected DF-1 cells (MOI = 1) was examined at 0, 2, 4, 6, 8 and 10 h after infection by Western blotting using anti-VP4 mAb.
Mentions: VP4 was previously reported to form tubule-like structures within IBDV-infected chicken embryo fibroblast cells (CEFs)17, but it’s unclear if the intracellular formation of tubular VP4 is specific for certain type of cells. We characterized VP4 expression in various IBDV-infected cells, including primary CEFs, DF-1 cells, Vero cells, HEK293T cells (in vitro) and intrabursal lymphocytes in chicken (in vivo) by immunofluorescence assay (IFA) using anti-VP4 monoclonal antibody (mAb). Confocal microcopy revealed that VP4 assembles within the cytoplasm and nucleus of various IBDV-infected cells (Fig. 1a). Specifically, intranuclear VP4 assembled into varying sizes of fibrils among IBDV-infected different cells, cytoplasmic VP4 in IBDV-infected CEF, DF-1 cells and HEK293T cells mainly formed needle-like fibrils, while cytoplasmic VP4 in the IBDV-infected Vero cells and intrabursal lymphoid cells formed large mass-like aggregates. These data demonstrated that intracellular VP4 assembly is an intrinsic feature of IBDV infection which is independent of cell types.

Bottom Line: Interestingly, the last C-terminal hydrophobic and amyloidogenic stretch (238)YHLAMA(243) with two "aggregation-prone" alanine residues was found to be essential for its intracellular self-assembly.Importantly, the assembly of VP4 significantly reduced the cytotoxicity of protease activity in host cells which potentially prevent the premature cell death and facilitate viral replication.This study provides novel insights into the formation mechanism and biological functions of the Avibirnavirus protease-related fibrils.

View Article: PubMed Central - PubMed

Affiliation: Key Laboratory of Animal Virology of Ministry of Agriculture, Zhejiang University, Hangzhou 310058, PR China.

ABSTRACT
Unlike other viral protease, Avibirnavirus infectious bursal disease virus (IBDV)-encoded viral protease VP4 forms unusual intracellular tubule-like structures during viral infection. However, the formation mechanism and potential biological functions of intracellular VP4 tubules remain largely elusive. Here, we show that VP4 can assemble into tubules in diverse IBDV-infected cells. Dynamic analysis show that VP4 initiates the assembly at early stage of IBDV infection, and gradually assembles into larger size of fibrils within the cytoplasm and nucleus. Intracellular assembly of VP4 doesn't involve the host cytoskeleton, other IBDV-encoded viral proteins or vital subcellular organelles. Interestingly, the last C-terminal hydrophobic and amyloidogenic stretch (238)YHLAMA(243) with two "aggregation-prone" alanine residues was found to be essential for its intracellular self-assembly. The assembled VP4 fibrils show significantly low solubility, subsequently, the deposition of highly assembled VP4 structures ultimately deformed the host cytoskeleton and nucleus, which was potentially associated with IBDV lytic infection. Importantly, the assembly of VP4 significantly reduced the cytotoxicity of protease activity in host cells which potentially prevent the premature cell death and facilitate viral replication. This study provides novel insights into the formation mechanism and biological functions of the Avibirnavirus protease-related fibrils.

No MeSH data available.


Related in: MedlinePlus