Endocytic Pathway Of Unidentified Receptor-binding Foot-and-mouth Disease Virus Entry Into Cells

Foot-and-mouth disease(FMD) is a highly contagious disease of domestic and wild cloven-hoofed animals, which is caused by foot-and-mouth disease virus(FMDV). FMDV is a small, no enveloped, positive-strand RNA virus belonging to the genus Aphthovirus within the family Picornaviridae. It contains about 8400 nucleotides. FMDV includes seven serotypes: O, A, C, Asia 1, and South African Territories(SAT) 1, 2 and 3, and includes several variants. The genetic and antigenic variability of FMDV endows the virus with a high potential for adaptation. It brings a series of difficult and complicated problem for controlling and eliminating the disease.Virus initialize the infection through receptor binding and cell receptors on the surface, being internalized to swallow in the vesicle, and then is transferred to the nucleus by unknown mechanisms. FMDV utilizes four members of the integrins, αvβ3, αvβ6, αvβ1 and αvβ8, as receptors, via a conserved Arg-Gly-Asp sequence in the G–H loop of VP1 that is the component of virus capsid. Some FMDV strains possess the ability to utilize heparn sulfate(HS) and alternative membrane molecules as receptors when adapted to tissue culture. Integrin-binding FMDV utilizes a clathrin-dependent mechanism to infect cells, and it is trafficked through early endosomes into recycling endosomes, HS-binding FMDV variants are internalized via caveola-mediated endocytosis. Low p H of endocytic organelles is important for the entry of most viruses. The acidic environment activates the penetration reactions, and allows viruses escape to the cytoplasm at specific locations. Then, the viruses are delivered to the hydrolytic lysosomes. Additionally, dynamin plays an important role in invasion of many viruses, which is essential for cellular membrane fission during vesicle formation, so it is required for clathrin- and caveola-mediated endocytosis.In this study, we choiced Chinese hamster ovary cell line(CHO-677) as a model, which lacks all of the known integrin receptors for FMDV and heparan sulfate(HS). Here, we evaluated the virus used unknown receptors internalization mechanism using various chemical inhibitors, dominant negative mutants or small interfering RNA(si RNA) to block different endocytic pathways. Firstly, The biological characteristics of FMDV invasion of host cell were analyzed. Then, dominant negative mutants and small interfering RNA(si RNA) were used to uggest that clathrin- and caveola-mediated endocytotic pathways are important during entry of virus into CHO-677 cells, and the results were validated by chemical inhibitors. In addition, this virus endocytosis into CHO-677 cells is dependent on dynamin and low-p H environment.In summary, our data indicate that FMDV Asia1/HN/CHA/06 enters CHO-677 cells by clathrin- and caveola-mediated endocytic pathways, and the entry is dependent on dynamin activity and a low-p H environment. It is possible that FMDV Asia1/HN/CHA/06 uses multiple entry pathways involving some unidentified receptors of FMDV. These findings might provide new ideas of FMDV study.