DISRUPTIONS IN GOLGI STRUCTURE AND MEMBRANE TRAFFIC IN A CONDITIONAL-LETHAL MAMMALIAN-CELL MUTANT ARE CORRECTED BY EPSILON-COP

The CHO cell temperature-sensitive mutant ldlF exhibits two defects in membrane traffic at the nonpermissive temperature (39.5 degrees C): r apid degradation of LDL receptors, possibly caused by endocytic missor ting, and disruption of ER-through-Golgi transport. Here, we show that at 39.5 degrees C, the Golgi in ldlF cells dissociated into vesicles and tubules. This dissociation was inhibited by A1F(4)(-), suggesting trimeric G proteins are involved in the dissociation mechanism. This r esembled the effects of brefeldin A on wild-type cells. We isolated a hamster cDNA that specifically corrected the ts defects of ldlF cells, but not those of other similar ts mutants (ldlE, ldlG, ldlH, and End4 ). Its predicted protein sequence is conserved in humans, rice, Arabid opsis, and Caenorhabditis elegans, and is virtually identical to that of bovine epsilon-COP, a component of the coatomer complex implicated in membrane transport. This provides the first genetic evidence that c oatomers in animal cells can play a role both in maintaining Golgi str ucture and in mediating ER-through-Golgi transport, and can influence normal endocytic recycling of LDL receptors. Thus, along with biochemi cal and yeast genetics methods, mammalian somatic cell mutants can pro vide powerful tools for the elucidation of the mechanisms underlying i ntracellular membrane traffic.