We isolated a novel yeast alpha-COP mutant, ret1-3, in which alpha-COP is degraded after cells are shifted to a restrictive temperature. ret1-3 cells cease growth at 28 degrees C and accumulate the ER precursor of carboxypeptidase Y (p1 CPY). In a screen for high copy suppressors of these defects, we isolated the previously unidentified yeast epsilon-COP gene. epsilon-COP (Sec28p) overproduction suppresses the defects of ret1-3 cells up to 34 degrees C, through stabilizing levels of alpha-COP. Surprisingly, cells lacking epsilon-COP (sec28 Delta) grow well up to 34 degrees C and display normal trafficking of carboxypeptidase Y and KKXX-tagged proteins at a permissive temperature. epsilon-COP is thus non-essential for yeast cell growth, but sec28 Delta cells are thermosensitive. In sec28 Delta cells shifted to 37 degrees C, wild-type alpha-COP (Ret1p) levels diminish rapidly and cells accumulate p1 CPY; these defects can be suppressed by alpha-COP overproduction. Mutant coatomer from sec28 Delta cells behaves as an unusually large protein complex in gel filtration experiments. The sec28 Delta mutation displays allele-specific synthetic-lethal interactions with alpha-COP mutations: sec28 Delta ret1-3 double mutants are unviable at all temperatures, whereas sec28 Delta ret1-1 double mutants grow well up to 30 degrees C. Our results suggest that a function of epsilon-COP is to stabilize alpha-COP and the coatomer complex.

• PMID: 9463377
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Journal Article | Research Support, Non-U.S. Gov't

Authors

Duden R, Kajikawa L, Wuestehube L, Schekman R

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