Folimycin(Concanamycin A), a Specific Inhibitor of V-ATPase, Blocks Intracellular Translocation of the Glycoprotein of Vesicular Stomatitis Virus before Arrival to the Golgi Apparatus.

Abstract

Key words: folimycin/concanamycinA/intracellular translocation of glycoproteins/vacuolar-type ATPase/vesicular stomatitis virus/the Golgi apparatus ABSTRACT. Folimycin (concanamycin A) specifically inhibited vacuolar-type ATPase as far as examined. Folimycin blocked excretion of the glycoprotein (G protein) of vesicular stomatitis virus into the medium and, instead, G protein was accumulated intracellularly.The intracellularly accumulated G protein electrophoresed a little faster than mature one. The N-glycan of the G protein was endoglycosidase H-sensitive, and terminal galactose and N-acetylglucosamine were not detected essentially on sequential digestion with exoglycosidases, indicating that processings known to occur in the Golgi apparatus do not take place in the presence of folimycin. The oligosaccharide chain of the G protein was determined to have a composition of Man8GlcNAc2 as analyzed by Bio-Gel P-4 column chromatography and high-performance liquid chromatography following digestion with a-and then with /5-mannosidase. Activities of mannosidase I and glycosyltransferases prepared from baby hamster kidney cells were not inhibited as far as examined, indicating that the incompleteness of the N-glycosidic chain in folimycin-treated cells is not caused by inhibition of processing enzymes. Taken together these observations suggest that folimycin blocks the intracellular translocation of G protein before the step of trimming by mannosidase I which is confined to the cis compartment of the Golgi. The intracellular localization of G protein as revealed by fluorescence microscopy was in good accordance with this assumption.

Prodigiosin 25‐C uncouples vacuolar type H+‐ATPase, inhibits vacuolar acidification and affects glycoprotein processing

“…Monensin blocks or slows down the intracellular transport of glycoprotein [21][22][23] and arrests the transport of VSV G protein prior to reaching the trans Golgi compartment. Bafilomycin A1 and concanamycin A also effectively block the cell surface expression of VSV G protein [9,10] but concanamycin A stops the transport of G protein from the endoplasmic reticulum to the cis Golgi compartment [9]. Compared with these inhibitors, the action of prodigiosin 25-C on the transport of VSV G protein appears to be quite leaky.…”

“…Preparation of VSV (New Jersey serotype) and labeling of VSV G protein were basically performed according to the methods described previously [9]. After BHK cells were absorbed with VSV (MOI 10) for t3 2.…”

“…V-ATPases have been identified and purified from a variety of organelles of eukaryotic cells and are clearly distinguished from other H ÷-ATPases by the inhibitor specificity [1,2]. Recently, bafilomycins [3] and concanamycins [4], originally found in microbial metabolites, have been shown to be specific inhibitors of VATPase and have become the most useful probes to study vacuolar acidification by V-ATPase [5][6][7][8][9][10][11].…”

“…bafilomycin B~ and concanamycin A), and Golgi mannosidase II inhibitor (swainsonine), were also more cytotoxic to those cells after addition of Con A [20]. These reagents, except for swainsonine, raise the pH in the acidic compartments and affect intracellular transport of glycoprotein [9,10,[21][22][23]. In this paper, we have studied the effect of prodigiosin 25-C on vacuolar acidification, intracellular transport of glycoprotein, and morphology of BHK cells, and we have found that prodigiosin 25-C uncouples V-ATPase and raises the pH of vacuolar organelles in intact cells.…”

Intracellular pH regulation in human Sertoli cells: role of membrane transporters

“…1; Table 1). Concanamycin A (10 mM), a specific inhibitor of the V-type ATPases, used at concentrations from 1 to 5 mM (LD 50 (mouse/oral) Z21 mg/Kg; IC 50 Z0.2-0.7 mM; Muroi et al 1993, Huss et al 2002, Oliveira et al 2004, caused a very small, although significant, decrease in the pHi recovery extension, as can be seen in Fig. 1 and Table 1.…”