Effect of N-glycosylation inhibition on the synthesis and processing of classical swine fever virus glycoproteins.
Classical swine fever virus (CSFV) is often used as a surrogate model in molecular studies of the closely related hepatitis C virus. In this report we have examined the effect of the inhibition of glycosylation on the survival and maturation of CSFV. Viral glycoproteins (E(rns), E1, E2) form biologically active complexes - homo- and heterodimers, which are indispensable for viral life cycle. Those complexes are highly N-glycosylated. We studied the influence of N-glycosylation on dimer formation using E(rns) and E2 glycoproteins produced in insect cells after infection with recombinant baculoviruses. The glycoproteins were efficiently synthesized in insect cells, had similar molecular masses and formed dimers like their natural counterparts. Surprisingly, the addition of tunicamycin (an antibiotic which blocks early steps of glycosylation) to insect cell culture blocked not only dimer formation but it also led to an almost complete disappearance of E2 even in monomeric form. Tunicamycin did not exert a similar effect on the synthesis and formation of E(rns) dimers; the dimers were still formed, which suggests that E(rns) glycan chains are not necessary for dimer formation. We have also found that very low doses of tunicamycin (much lower than those used for blocking N-glycosylation) drastically reduced CSFV spread in SK6 (swine kidney) cell culture and the virus yield. These facts indicate that N-glycosylation inhibitors structurally similar to tunicamycin may be potential therapeutics for the inhibition of the spread of CSFV and related viruses.
Tyborowska
Jolanta
J
Department of Molecular Virology, Intercollegiate Faculty of Biotechnology, University of Gdansk, and Medical University of Gdansk, Gdańsk, Poland. jtybor@biotech.ug.gda.pl
Zdunek
Ewa
E
Szewczyk
Bogusław
B
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D015153:Blotting, Western; D002460:Cell Line; D006692:Classical Swine Fever Virus / Q000378:metabolism*; D019084:Fluorescent Antibody Technique, Indirect; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D006031:Glycosylation; D018613:Microscopy, Confocal; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D013552:Swine; D014415:Tunicamycin / Q000494:pharmacology; D014764:Viral Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
D006023:Glycoproteins; D011500:Protein Synthesis Inhibitors; D014764:Viral Proteins; D014415:Tunicamycin