Full length articleRoles of extracellular matrix components in Tiger frog virus attachment to fathead minnow (Pimephales promelas) cells
Introduction
Extracellular matrix (ECM) is a non-cellular macromolecular network composed of collagens, glycosaminoglycans (GAGs), fibronectin, laminin, hyaluronic acid, and several other glycoproteins [1]. Matrix components bind each other to transduce signals into cells from the ECM, which regulates diverse cellular functions, such as survival, growth, migration, and differentiation, and are also vital for many virus infections [2].
GAGs play important roles in virus attachment, usually as a virus attachment factor in mammals [3]. Heparan sulfate is one of the most common GAGs in mammals; and it involved in cell attachment of many viruses, such as Herpes simplex virus, Respiratory syncytial virus, Hepatitis B virus, and Dengue virus [[4], [5], [6], [7]]. ECM proteins are also involved in the attachment of many viruses onto host cells, for example, Human T-cell leukemia virus type 1 hijacking collagen IV as an anchor to tether and enhance viral infection transmission [8]. Human papillomavirus virions bind to laminin on the ECM and then transfer to interact with their entry receptors in keratinocytes [9]. Increasing research showed that ECM components also play important roles in the attachment step of aquatic animal viruses. Cell surface heparan sulfate is an important factor for the binding of Andrias davidianus ranavirus and Rana grylio virus to target cells [10]. The attachment and entry of Infectious hematopoietic necrosis virus in zebrafish is mediated by fibronectin [11]. Viral hemorrhagic septicemia virus employs fibronectin as attachment factor in rainbow trout (Oncorhynchus mykiss) gonad cells because anti-fibronectin antibody can block infection; purified fibronectin also binds specifically to virions [12]. However, the studies on the roles of ECM components in the aquatic virus attachment remain limited.
Iridoviruses are large, icosahedral cytoplasmic dsDNA virus, and measure 120–350 nm in diameter [13]. Iridoviridae is divided into two subfamilies and six genera: subfamily Alpha-iridoviridae includes Lymphocystivirus, Megalocytivirus, and Ranavirus; subfamily Beta-iridoviridae includes Chloriridovirus, Decapodiridovirus and Iridovirus [15,16]. Tiger frog virus (TFV) belongs to the genus Ranavirus, and it has been isolated from diseased tiger frog Hoplobatrachus tigerinus [[14], [17]]. TFV causes great losses of commercial field in Southern China; the estimated mortality of cultured tadpoles reaches about 95%; similar to farms, TFV also causes noticeable threat to natural populations of fish and amphibians [17]. TFV is the first reported complete genome sequence in the genus Ranavirus; its genome comprises double-stranded DNA of 105,057 base pairs in length and is organized by 105 non-overlapping open reading frames, sharing marked sequence identity with Frog virus 3, the type species of the genus [[14], [16]]. Recently, the mechanisms of TFV entry, immune escape, and release have been identified; these mechanisms include TFV uptake into HepG2 cells via a pH-dependent atypical caveola-mediated endocytosis pathway [18]. The molecular mechanism of TFV regulated anti-apoptosis occurs through the interaction of TFV ORF104R with the voltage-dependent anion-selective channel protein 2 protein [19]. TFV VP080L could negate the function of cellular lipopolysaccharide-induced tumor necrosis factor-alpha factor in endosomal sorting complex required for transport (ESCRT) to impair endosomal sorting and trafficking [20]. TFV relies on the network of the cellular ESCRT proteins to complete the process of virus budding [21]. However, attachment as the first step of TFV infection is still unclear.
In the present study, the roles of the primary components of ECM in the TFV attachment were investigated.
Section snippets
Cells and viruses
Fathead minnow (Pimephales promelas, FHM) cells were maintained in M199 medium (Gibco, USA) supplemented with 10% fetal bovine serum (FBS, Gibco, USA) at 27 °C and 5% CO2. TFV was originally isolated from diseased tiger frog tadpoles in Nanhai, Guangdong, China and was maintained by our laboratory [19]. TFV was propagated in FHM cells and purified as described previously [21]. Briefly, cells were infected with the virus with a multiplicity of infection of 0.1 for 6 days. After freeze-thawing
TFV attachment is a relatively rapid process
The kinetics of virus attachment were investigated by detection of binding virus in the cells surface at different times to understand the process of TFV attachment onto host cells. The levels of TFV MCP and mcp gene represented the levels of viral protein and viral genome DNA copies, which were detected by Western blot and qPCR analyses. As shown in Fig. 1A–B, TFV MCP and viral genome DNA copies significantly increased after virus incubation with cells at 10 min, indicating that TFV
Discussion
Viral attachment is important for the first step of infection to ensure that viruses remain on the cell surface. Viruses attach to permissible cell with non-strict-specificity, followed by further specific receptor–ligand interaction to enter cells [23,24]. At the attachment step, a virus might come into contact with the ECM components, which almost or completely surround the cell [25]. Most of the work on viral attachment to cell surface factors has been conducted in vitro, and the main
CRediT authorship contribution statement
Chang Liu: performed the experiments, wrote the article. All authors read and approved the final manuscript. Jian He: and. Shao-Ping Weng: and. Jian-Guo He: conceived the study, performed sampling, data, and laboratory analyses. Chang-Jun Guo: and, and.
Declaration of competing interest
The authors declare no competing interests.
Acknowledgement
This work was supported by the National Key Research and Development Program of China, China (Nos. 2018YFD0900504 and 2018YFD0900501), the National Natural Science Foundation of China, China (No. 31702381), the Science and Technology Planning Project of Guangzhou, China (Nos. 201904020043 and 201607020014), the China Agriculture Research System (No. CARS-46), the Guangdong Provincial KeyResearch and Development Program, China (No. 2019B020217001), and Guangdong Provincial Special Fund for
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2022, Fish and Shellfish ImmunologyCitation Excerpt :Extracellular matrix is a macromolecular network and mainly composed of glycosaminoglycans, fibronection, heparin sulfate, collagen, which played the important roles in aquatic animal virus infections [33,34]. The available reports proved that extracellular matrix components were essential for many aquatic viruses to attach and entry into the host, such as viral hemorrhagic septicemia virus [35], IHNV [36], tiger frog virus [34], andrias davidianus ranavirus and rana grylio virus [37]. Therefore, it was speculated that LTCONS_00075232 (zinc finger protein 384-like co-expressed gene) possibly involved in regulating the changes of the extracellular matrix in IHNV infected cells, thereby influencing viral invasion.