Tetraspanins as Potential Therapeutic Candidates for Targeting Flaviviruses

Abstract

Tetraspanin family of proteins participates in numerous fundamental signaling pathways involved in viral transmission, virus-specific immunity, and virus-mediated vesicular trafficking. Studies in the identification of novel therapeutic candidates and strategies to target West Nile virus, dengue and Zika viruses are highly warranted due to the failure in development of vaccines. Recent evidences have shown that the widely distributed tetraspanin proteins may provide a platform for the development of novel therapeutic approaches. In this review, we discuss the diversified and important functions of tetraspanins in exosome/extracellular vesicle biology, virus-host interactions, virus-mediated vesicular trafficking, modulation of immune mechanism(s), and their possible role(s) in host antiviral defense mechanism(s) through interactions with noncoding RNAs. We also highlight the role of tetraspanins in the development of novel therapeutics to target arthropod-borne flaviviral diseases.

Keywords: arthropods; exosomes; flaviviruses; ncRNAs; tetraspanins; transmission.

Figure 1

Tetraspanin structural organization and domain analysis from arthropod and vertebrate host. (A) Schematic diagram showing tetraspanin domain organization predicted at PROSITE (ExPASy) using primary amino acid sequence of mosquito A. aegypti Tsp29Fb (accession no: AAEL012532-RA), human- CD9 (accession no: NP_001760.1), CD63 (accession no: NP_001771.1), CD81 (accession no: NP_004347.1), and CD151 (accession no: NP_001034579.1). N-glycosylation sites are predicted using NetNGlycb (http://www.cbs.dtu.dk/services/NetNGlyc) and PROSITE (ExPASy). N-glycosylation sites, which were predicted in both web tools, are shown. Domain analysis demonstrates presence of conserved tetraspanin domain, 4 transmembrane regions, and variable number of glycosylation/myristoylation sites, and amino acid sequences. Different regions in tetraspanin proteins are indicated with different colors and labels. (B) Schematic representation of several characteristic features in tetraspanin protein is shown in the context of its transmembrane architecture. Tetraspanins have four highly conserved transmembrane domains (TM1-4), two extracellular portions known as small extracellular loop (EC1/SEL) and large extracellular loop (EC2/LEL), one intracellular loop (trans-passing from domain 2-3), and N- and C-terminal tails. EC2/LEL domain is conserved among several tetraspanins with 2–4 disulphide bonds (indicated with black lines) formed between cysteine residues in CCG-motif. These residues are binding sites for many interacting proteins and a strong epitope region for anti-tetraspanin antibodies. Numerous tetraspanins show glycosylation (shown in green lines) in the extracellular loops and palmitoylation sites (shown in blue line) at the intracellular border of the four transmembrane domains.

Figure 2

Proposed model for the role of tetraspanins in viral interactions. Mosquito tetraspanin Tsp29Fb is involved in DENV2/3 transmission from arthropod to mammalian cells, and Tsp C189 play a key role in cell-to-cell spreading of viral particles (indicated as step 1). Tetraspanin Tsp29Fb directly interacts with viral E-protein and perhaps might act as co-receptor for viral entry (denoted as step 2), and in specific serve as a receptor in case of arbovirus entry during clathrin-dependent receptor-mediated endocytosis (shown as step 3) and/or for internalization of viral particles by endocytosis (indicated as step 4). Tetraspanin proteins trigger the fusion and viral uncoating process (represented as step 5). Tetraspanins in association with several other host proteins are implicated in replication (denoted as step 6) and in translation process (shown as step 7). Tetraspanins participates to enable the delivery of viral genomes into the nucleus for successful infection and assembly processes (noted as steps 8). Tetraspanin contribution is highlighted in budding and assembly formation (noted as step 9) and in membrane fusion events can be exploited for viral exit from cells via direct release of secretory vesicles such as exosomes (represented as step 10). The step numbers have been classified into two color codes (light blue or dark gray). Role of arthropod tetraspanin in entry/exit of flaviviruses is proposed research and indicated in light blue color, whereas the published data is represented as dark gray circles.

Figure 3

Role of tetraspanins in exosome biogenesis and modulation of immune signaling. (A) Role of tetraspanins in exosome biogenesis is shown. The early endosome formation is the first step in vesicle uptake and recycling. Development of early multivesicular bodies (MVBs, shown in green complex) is dependent on sorting of cytosolic proteins, clathrin-coated membranes, RNA molecules, tetraspanin-enriched domains (TEMs), and lipid rafts. The sorting and packaging of several other molecules into exosomes cannot be ruled out and has not been shown for simplicity. Some mature multivesicular bodies (MVBs) fuse with the hydrolytic lysosome, where the vesicular cargo is subsequently degraded. The membranes of late MVBs fuses with the plasma membrane resulting in the release of exosomes/EVs into the extracellular environment. (B) Tetraspanins role in miRNA modulation is shown. Tetraspanins directly or indirectly interacts with different signaling molecules and receptor(s) at the membranes/lipid rafts, and organize into the specialized tetraspanin-enriched micro-domains (TEMs) that might play significant role during exosome cargo sorting (miRNAs, RNA, and proteins) from viral-infected cells to recipient cell. (C) Tetraspanin involvement in signaling pathway is shown. Exosomes are potential in the context of modulation of an immune response through their ability to present MHC–peptide complexes to specific cells, resulting in activation of T cells and antigen presentation cells.