INHIBITION OF ENDOPLASMIC RETICULUM-TO-GOLGI TRAFFIC BY POLIOVIRUS PROTEIN 3A - GENETIC AND ULTRASTRUCTURAL ANALYSIS

Poliovirus protein 3A, only 87 amino acids in length, is a potent inhi bitor of protein secretion in mammalian cells, blocking anterograde pr otein traffic from the endoplasmic reticulum (ER) to the Golgi complex . The function of viral protein 3A in blocking protein secretion is ex tremely sensitive to mutations near the N terminus of the protein, Del etion of the first 10 amino acids or insertion of a single amino acid between amino acids 15 and 16, a mutation that causes a cold-sensitive defect in poliovirus RNA replication, abrogates the inhibition of pro tein secretion although wild-type amounts of the mutant proteins are e xpressed. Immunofluorescence light microscopy and immunoelectron micro scopy demonstrate that 3A protein, expressed in the absence of other v iral proteins, colocalizes, with membranes derived from the ER. The pr ecise topology of 3A with respect to ER membranes is not known, but it is likely to be associated with the cytosolic surface of the ER. Alth ough the glycosylation of 3A in translation extracts has been reported , we show that tunicamycin, under conditions in which glycosylation of cellular proteins is inhibited, has no effect on poliovirus growth, T herefore, glycosylation of 3A plays no functional role in the viral re plicative cycle. Electron microscopy reveals that the ER dilates drama tically in the presence of 3A protein, The absence of accumulated vesi cles and the swelling of the ER-derived membranes argues that ER-to-Go lgi traffic is inhibited at the step of vesicle formation or budding f rom the ER.