January 23, 2019;
Class A Scavenger Receptors Are Used by Frog Virus 3 During Its Cellular Entry.
Frog virus 3 (FV3) is the type species of the genus Ranavirus (family Iridoviridae). FV3 and FV3-like viruses are globally distributed infectious agents with the capacity to replicate in three vertebrate classes (teleosts, amphibians, and reptiles). At the cellular level, FV3 and FV3-like viruses can infect cells from virtually all vertebrate classes. To date, the cellular receptors that are involved in the FV3 entry process are unknown. Class A scavenger receptors (SR-As) are a family of evolutionarily conserved cell-surface receptors that bind a wide range of chemically distinct polyanionic ligands and can function as cellular receptors for other DNA viruses, including vaccinia virus and herpes simplex virus. The present study aimed to determine whether SR-As are involved in FV3 cellular entry. By using well-defined SR-A competitive and non-competitive ligand-blocking assays and absolute qPCR, we demonstrated that the SR-A competitive ligands drastically reduced the quantities of cell-associated viral loads in frog cells. Moreover, inducing the expression of a human SR-AI in an SR-A null cell line significantly increased FV3⁻cell association. Together, our results indicate that SR-As are utilized by FV3 during the cellular entry process.
transport of virus in host, cell to cell
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Figure 1. Effects of SR-A competitive and non-competitive ligands on the cell-associated FV3 loads
in four tadpole cell lines. The American toad BufoTad cell line, bullfrog BullTad-leg cell line, green
frog GreenTad-HF2 cell line, and wood frog WoodTad-rpe cell line were pre-treated with 250 g/mL of
the SR-A competitive ligands (fucoidan, poly inosine (pI)) (A–D) or non-competitive ligands (fetuin,
poly cytosine (pC)) (E–H) for 30 min. FV3 was added at an multiplicity of infection MOI of 1.0 for 2 h.
The FV3 DNA viral loads were determined by absolute qPCR. Data are presented as means SEM
(n = 3). Statistical analysis using a one-way ANOVA test and Tukey’s multiple comparisons test with
95% confidence intervals was performed. p values less than 0.05 are considered statistically significant.
Groups with different letters are statistically different from one another.
Figure 2. Effects of SR-A competitive and non-competitive ligands on the cell-associated FV3 load in
X. laevis CSF-1 Ms and IL-34 Ms. The X. laevis CSF-1 Ms and IL-34 Ms were derived by culturing
adult frog bone marrow cells with recombinant CSF-1 or IL-34, respectively, for 5 days. The Ms
were then pre-treated with 200 g/mL of the SR-A competitive ligands (DxSO4, fucoidan) (A,B) and
non-competitive ligands (ChSO4, fetuin) (C,D) for 1 h and challenged with FV3 (MOI of 0.5) for 2 h.
The FV3 DNA viral loads were determined by absolute qPCR. The results are presented as means
SEM (n = 3–5). Statistical analysis using a one-way ANOVA test and Tukey’s multiple comparisons
test with 95% confidence intervals was performed. p values less than 0.05 are considered statistically
significant. Groups with different letters are statistically different from one another.
Figure 3. Effect of induced human SR-AI (hSR-AI) expression on the cell-associated FV3 loads in
SR-A-null A549 cells. hSR-AI expression was induced with 200 ng/mL doxycycline (DOX). Control
SR-A-null and DOX-treated hSR-AI-expressing cells were infected with FV3 at an MOI of 0.5 for 1 h.
Total DNA was subsequently harvested and subjected to absolute qPCR analyses for FV3 DNA loads.
Data are presented as means SEM (n = 3). Statistical analysis using a Student’s t-test withWelch’s
correction was performed. ***p < 0.001.