Effect of caffeine and reduced temperature (20 degrees C) on the organization of the pre-Golgi and the Golgi stack membranes

Localization of marker proteins of the pre-Golgi membranes and the Golgi stack demonstrated an interesting, differential effect of caffeine on the distribution of these membrane compartments. The p58-positive pre-and cis-Golgi membranes lost their normal perinuclear localization while the localization of a Golgi stack protein, man II, remained unaffected. The normal distribution of p58 was restored when caffeine was removed. When the localization of p58 was studied with immunoelectron microscopy in caffeine-treated cells at 20~ the typical tubulovesicular p58-positive elements (Saraste and Svensson, 1991) were absent from the Golgi region and p58 was found only to be localized in a few small vesicles associated with the clusters of vesicles in the Golgi region. Even though immunoperoxidase technique is by no means a quantitative method, when compared with the control cells, the number of p58-positive membrane elements in the Golgi region seemed to be lower in caffeine-treated cells at reduced temperature. The statement that p58 is translocated away from the perinuclear 1o-calization in caffeine-treated cells is based on the results obtained in the immunofluorescence experiments, where distribution of the bulk of p58 can be observed. Overall, the immunoelectron microscopic finding is in agreement with the immunofluorescence result in Fig. 5 a. As suggested by the immunofluorescence experiments, the bulk of the p58 seems to be translocated away from the perinuclear localization in caffeine-treated cells. These resuits suggest that caffeine and reduced temperature are able to induce a selective retrograde translocation of pre-and cis-Golgi membranes while the localization of the Golgi stack membranes remains unaffected. This was further confirmed by the unaffected localization of/~-COP protein. Possible explanation to the observed loss of the perinuclear labeling of p58 could be that in the presence of caffeine the pre-and cis-Golgi elements are induced to move toward the periphery of the cell. Another possibility is that as the membrane traffic from the ER is inhibited and at least the BFA-induced retrograde movement of membranes is not inhibited, p58 could finally be recycled to the ER, being unable to exit from there. Whatever the solution turns out to be, caffeine seems to make it possible to manipulate the membranes operating in the ER-to-Golgi traffic in a novel way. Interestingly, Lewis and Pelham (1992) have recently shown that, by overexpressing the ligand for the KDEL receptor, the bulk of the receptor is translocated to the ER.