Hepatitis B surface antigen assembles in a post-ER, pre-Golgi compartment

The observation that oligomer crosslinks only form after exit from the ER suggests that there must be something different about the later compartment which allows these oligomer crosslinks to form. The obvious candidate for this difference is the absence of PDI. We tested the effect of PDI Figure 7 . The time course of HBsAg secretion from SV24 cells. The autoradiograph inA shows the secreted material which was immunoprecipitated from the medium after a 10-rain radioactive pulse and the indicated chase times. B shows a quantitative analysis of the secretion. The X indicates the immunoprecipitable counts released into the medium as a function of chase time with the nonspecific specifically bound counts subtracted. The fitted curve (----) shows the equation 16000 cpm* (1-exp(-t/2.5h)) which was the best weighted least squares fit to the data. The corresponding half-time for the appearance of HBsAg particles in the culture medium is •2 h. The ratios of glycosylated monomers to the nonglycosylated monomers in the particles secreted as a function of chase time (~) were measured from the autoradiograph in A. The average ratio of glycosylated HBsAg to nonglycosylated HBsAg (gp/p) was 0.551 and is indicated by the horizontal line. on the oligomer crosslinks by incubating secreted, fully crosslinked HBsAg particles with a lysate of Sf9 cells which had been infected with a rat PDI-cDNA bearing recombinant baculovirus and so expressed high levels of this protein (Fig. 9) . As a control, lysate from cells infected with wild type baculovirus was used. The effect of increasing amounts of the PDI-containing lysate is the conversion of the oligomeric forms of HBsAg to the dimeric form ( Fig. 9 C) . The level of monomer remains approximately the same. Although secreted HBsAg contains no monomer, the prolonged incubation with glutathione causes the direct conversion of a small fraction of the oligomer to monomer. This can also be observed in incubations in which no cell lysate has been added (data not shown). This experiment shows that the oligomer crosslinks of HBsAg particles are not stable in the presence of PDI although the dimer crosslink formation is promoted.