EPSILON-COP IS A STRUCTURAL COMPONENT OF COATOMER THAT FUNCTIONS TO STABILIZE ALPHA-COP

We isolated a novel yeast alpha-COP mutant, ret1-3, in which alpha-COP is degraded after cells are shifted to a restrictive temperature, ret 1-3 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, me isolated the previously unidentified yeast epsil on-COP gene. epsilon-COP (Sec28p) overproduction suppresses the defect s of ret1-3 cells up to 34 degrees C, through stabilizing levels of al pha-COP. Surprisingly, cells lacking epsilon-COP (sec28 Delta) grow we ll up to 34 degrees C and display normal trafficking of carboxypeptida se Y and KKXX-tagged proteins at a permissive temperature, epsilon-COP is thus non-essential for yeast cell growth, but sec28 Delta cells ar e 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. Muta nt coatomer from sec28 Delta cells behaves as an unusually large prote in complex in gel filtration experiments, The sec28 Delta mutation dis plays allele-specific synthetic-lethal interactions with alpha-COP mut ations: sec28 Delta ret1-3 double mutants are unviable at all temperat ures, whereas sec28 Delta ret1-1 double mutants grow well up to 30 deg rees C, Our results suggest that a function of epsilon-COP is to stabi lize alpha-COP and the coatomer complex.