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

Abstract

Abstract. In the present study we have dissected the transport pathways between the ER and the Golgi complex using a recently introduced (Kuismanen, E., J. Jiintti, V. Miikiranta, and M. Sariola. 1992. J. Cell Sci. 102:505-513) inhibition of transport by caffeine at 20~ Recovery of the Golgi complex from brefeldin A (BFA) treatment was inhibited by caffeine at reduced temperature (20~ suggesting that caffeine inhibits the membrane traffic between the ER and the Golgi complex. Caffeine at 20~ did not inhibit the BFA-induced retrograde movement of the Golgi membranes. Further, incubation of the cells in 10 mM caffeine at 20~ had profound effects on the distribution and the organization of the pre-Golgi and the Golgi stack membranes. Caffeine treatment at 20~ resulted in a selective and reversible translocation of the pre-and cis-Golgi marker protein (p58) to the periphery of the cell. This caffeine-induced effect on the Golgi complex was different from that induced by BFA, since mannosidase II, a Golgi stack marker, remained perinucleady located and the Golgi stack coat protein, B-COP, was not detached from Golgi membranes in the presence of 10 mM caffeine at 20~ Electron microscopic analysis showed that, in the presence of caffeine at 20~ the morphology of the Golgi stack was altered and accumulation of numerous small vesicles in the Golgi region was observed. The results in the present study suggest that caffeine at reduced temperature (20~ reveals a functional interface between the pre-Golgi and the Golgi stack.

Immunocytochemical analysis of Uukuniemi virus budding compartments: role of the intermediate compartment and the Golgi stack in virus maturation

“…After fixation the cells were washed with DBS buffer (phosphate-buffered saline supplemented with 0.2% bovine serum albumin and 0.05% saponin, pH 7.2) followed by treatment with 0.05 M lysine in DBS buffer to quench free aldehyde groups. The cells were labeled with rabbit antinucleocapsid antibodies and peroxidase-conjugated anti-rabbit F(ab) 2 secondary antibodies (Immunotech, Marseille, France) as described previously (11,17). For labeling with antibodies to the p58 protein and mannosidase II (ManII), the cells were fixed with the periodate-lysine-paraformaldehyde fixative (3,23).…”

Molecular chaperones involved in protein degradation in the endoplasmic reticulum: quantitative interaction of the heat shock cognate protein BiP with partially folded immunoglobulin light chains that are degraded in the endoplasmic reticulum.

“…However, this would presumably involve a vesicular transport step which is excluded by the results of temperature-sensitivity experiments reported here. In addition, caffeine, which blocks transport from the ER to the intermediate compartment (28), had no effect on Lchain degradation (M.R.K, unpublished data).…”

Golgi Apparatus Immunolocalization of Endomannosidase Suggests Post-Endoplasmic Reticulum Glucose Trimming: Implications for Quality Control

“…After 60 min at 37°C the inherent endomannosidase immunofluorescence pattern was observed. Jäntti and Kuismanen (1993) and Jäntti et al (1997) reported that a Golgi protein and intermediate compartment proteins segregate after brefeldin A redistribution at the level of the 15°C peripheral sites. When subsequently exposed to caffeine at 20°C, p58 remained in the peripheral sites, whereas Golgi mannosidase II became centralized perinuclearly.…”