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Characterization of SA-11 rotavirus receptorial structures on human colon carcinoma cell line HT-29
Abstract:
The involvement of different cell membrane components in the receptor structures for SA-11 rotavirus was investigated. As experimental model, the human enterocyte-like HT-29 cell line, was used because of its closer resemblance to the in vivo viral cellular target as compared to other in vitro systems. Rotavirus was incubated with whole membranes or their separated protein and lipid fractions before infection. Either isolated cell membranes or lipid components were capable of binding to the virus and to prevent infection, whereas proteins did not show any inhibitory activity. Among lipids, the glycolipid fraction was shown to impede rotaviral antigen synthesis with a dose-dependent relationship, whereas phospholipids failed to prevent viral infection. To confirm these findings, membranes and target cells were subjected to different enzymatic treatments prior to infection. In addition, HT-29 cells were also incubated with different lectins before infection. The blocking activity of membranes was inhibited by treatment with ceramide glycanase, neuraminidase, and beta-galactosidase but not by treatment with proteases or heat (100 degrees C). Viral infection was prevented by preincubation of target cells with lectins specific for sialic acid and galactose or with ceramide glycanase, neuraminidase, and beta-galactosidase, whereas protease treatments were not active. The results of these experimental procedures indicate that glycolipids containing specific carbohydrate moieties, such as sialic acid and galactose, contribute to the SA-11 rotavirus receptor structure on HT-29 cells.
Insights
Rotavirus uses specific glycolipids on host cell membranes as receptors. These glycolipids, containing sialic acid and galactose, are crucial for viral entry and infection.
Area of Science:
- Virology
- Cell Biology
- Glycobiology
Background:
- Rotaviruses are a leading cause of diarrheal disease in infants.
- Understanding rotavirus-host cell interactions is crucial for developing antiviral strategies.
- The human enterocyte-like HT-29 cell line serves as a relevant model for studying rotavirus infection.
Purpose of the Study:
- To investigate the role of cell membrane components in SA-11 rotavirus binding and infection.
- To identify specific lipids or proteins involved in rotavirus receptor structures.
- To elucidate the carbohydrate moieties essential for rotavirus-host cell recognition.
Main Methods:
- Incubation of rotavirus with whole cell membranes, lipid, and protein fractions of HT-29 cells.
- Enzymatic treatments (ceramide glycanase, neuraminidase, beta-galactosidase, proteases) of membranes and cells.
- Preincubation of HT-29 cells with specific lectins.
- Assessment of viral binding and antigen synthesis post-incubation.
Main Results:
- Lipid components, particularly glycolipids, effectively bound SA-11 rotavirus and inhibited infection.
- Proteins did not exhibit inhibitory activity against rotavirus infection.
- Enzymatic removal of sialic acid and galactose residues from cell membranes blocked viral binding.
- Lectins specific for sialic acid and galactose prevented viral infection.
Conclusions:
- Glycolipids containing sialic acid and galactose are essential components of the SA-11 rotavirus receptor on HT-29 cells.
- These specific carbohydrate structures mediate the interaction between rotavirus and host cells.
- The findings provide insights into the molecular mechanisms of rotavirus entry.