Internalization and trafficking mechanisms of coxsackievirus B3 in HeLa cells

Abstract

Coxsackievirus B3 (CVB3) is nonenveloped and has a single-stranded positive-sense RNA genome. CVB3 induces myocarditis and ultimately dilated cardiomyopathy. Although there are mounting evidences of an interaction between CVB3 particles and the cellular receptors, coxsackievirus and adenovirus receptor (CAR) and decay-accelerating factor (DAF), very little is known about the mechanisms of internalization and trafficking. In the present study, we used the CVB3 H3 strain, which is CAR-dependent but DAF-independent Woodruff variant and found that during entry, CVB3 particles were colocalized in clathrin, after interacting primarily with CAR, which was not recycled to the plasma membrane. We also found that CVB3 internalization was dependent on the function of dynamin, a large GTPase that has an essential role in endocytosis. Heat-shock cognate protein, Hsc70, which acts as a chaperone in the release of coat proteins from clathrin-coated vesicles (CCV), played a role in CVB3 trafficking processes. Moreover, endosomal acidification was crucial for CVB3 endocytosis. Finally, CVB3 was colocalized in early endosome autoantigen 1 (EEA1) molecules, which are involved in endosome-endosome tethering and fusion. In conclusion, these data together indicate that CVB3 uses clathrin-mediated endocytosis and is transcytosed to early endosomes.

ERK MAP kinase-activated Arf6 trafficking directs coxsackievirus type B3 into an unproductive compartment during virus host-cell entry

“…Once in the circulation, CVB3 can infect a myriad of cell types, including those in the gut, liver, pancreas and heart. Nonpolarized cells can be infected, and virus entry into these cells may occur via clathrin-coated pits and the early endosome network (Chung et al, 2005).…”

Toward Development of Artificial Viruses for Gene Therapy: A Comparative Evaluation of Viral and Non‐viral Transfection

“…Direct penetration, or fusion, is thought to be reserved for nonenveloped viruses, although evidence is accumulating that even non-enveloped viruses enter cells through the same endocytic routes as enveloped viruses. 34,39 The endocytic pathways vary depending on the virus and are reported to include clathrin-dependent 37,39,40 and caveolin-dependent 43 routes, as well as macropinocytosis 48 ( Table 2). Clathrin-mediated endocytosis is the predominant entry mechanism for viruses, including hepatitis B and C, coxsackievirus B3, and AAV (adeno-associated virus).…”

“…48 Clearly, as with non-viral vectors, viruses mediate entry through a variety of routes, with vectors initiating different routes in various cell lines. 22,39,65 Interestingly, despite their evolutionary advantage over non-viral vectors, viruses may not demonstrate better cell penetration abilities. 66 Studies demonstrate that co-incubation with lipoplexes improves viral internalization, presumably due to the nonspecific interactions between cells and lipoplexes, rather than the receptormediated interactions required for viral internalization.…”

“…Many viruses, including AAV, Ad, coxsackievirus B, Semliki Forest, hepatitis C, and influenza, require trafficking to and acidification of the late endosomal compartment for effective transduction. 34,35,37,39,40,[77][78][79] It is thought that processing of the viral capsid in an acidic environment may be necessary to activate viral components required for downstream processing, such as nuclear translocation. 77,78 Cell-line transduction dependence also correlates with intracellular trafficking: AAV viruses trafficked through the endo-lysosomal pathway mediate transduction in 293 cells; in contrast, they escape to the cytosol prior to acidification in NIH 3T3 cells, presumably through a different pathway, and are transfection incompetent.…”

Coxsackievirus B3 used as a gene therapy vector to express functional FGF2

“…13 CVB3 enters its target cells through the coxsackievirus and adenovirus receptor (the main receptor) and the membrane protein decay accelerating factor (a coreceptor). 14,15 Recently, several studies have reported that CVB3 can be used as a vector to express foreign genes, such as hepatitis C virus E2 and some cytokines. 16 --18 However, no recombinant CVB3 vector has yet been used in a disease model.…”