BREFELDIN-A SENSITIVITY AND RESISTANCE IN SCHIZOSACCHAROMYCES-POMBE -ISOLATION OF MULTIPLE GENES CONFERRING RESISTANCE

The fungal metabolite brefeldin A (BFA) causes the inhibition of prote in Secretion and the disruption of the structure and function of the G els complex in mammalian cells, Here we show that BFA has identical ef fects in the fission yeast Schizosaccharomyces pombe which normally co ntains a Golgi complex of stacked cisternae similar to the Golgi compl exes in animal cells. After treatment with BFA, secretion was inhibite d, Golgi complexes disappeared, and there was an accumulation of endop lasmic reticulum. These results indicate that the effects of BFA in fu ngi are very similar to those in mammalian cells and provide direct ev idence for an effect of BFA on Golgi morphology in fungi. Five spontan eous BFA-resistant mutants were isolated. Genetic analysis showed that the mutations conferring BFA resistance were dominant and in two sepa rate linkage groups. One of the BFA-resistant mutations was found to b e allelic: to crm1, a gene affecting chromatin structure. Ah BFA-resis tant mutants overexpressed a 20-kDa protein, and the corresponding gen e obr1 was isolated and sequenced. However, obr1 overexpression was no t sufficient to confer BFA resistance. Plasmids capable of conferring BFA resistance to wild type cells were isolated from libraries constru cted from the two BFA-resistant mutants. These plasmids contain six di fferent genes capable of conferring resistance when present in high co py. One of these genes encoded the transcription factor pap1, a homolo g of the mammalian AP1 protein. The overexpression of pap1 probably co nfers BFA resistance indirectly by inducing expression of one or more other proteins. The isolation of several genes conferring BFA resistan ce suggests several mechanisms are involved.