Figure 4. Mutants with con- ditional defects in vesicular transport between the ER and Golgi complex have al- tered ER structure at non- permissive temperature. Im- ages of cells expressing Sec63-GFP acquired while focusing on either the center or periphery of the cells. Bars, 5 mm. Arrows point to daughter cells in some cells. A, Images of sec27-95 cells grown either at 258C or 3 h after shift to nonpermissive temperature (378C). B, Im- ages of sec21-1, sec23-1, and ret1-3 cells 3 h after shift to 378C. C, Images of ufe1-1 cells 2 h after shift to 378C and cdc48-3 cells 4 h after shift to 378C. tional defects in other components of COPI (sec21-1 and ret1-3), a mutant with a defect in the functionally related coat complex COPII (sec23-1), and Ufe1p, an ER-local- ized vesicle soluble N-ethylmaleimide–protein attach- ment protein receptor (v-SNARE; ufe1-1; Fig. 4, B and C). Mutants with defects in other components of COPI or COPII (sec28D::HIS3, sec27-1, sec13-1, and sec16-2) had disorganized ER structure at nonpermissive temperature, but did not have the dramatic reduction in the amount of ER in daughter cells seen in the other mutants (not shown). It is unclear why this phenotype is seen in some mutants with defects in ER–Golgi complex traffic and not in others. Perhaps the rate and extent of inactivation of the mutant proteins differs in these two groups. This might also explain why this phenotype is seen in the sec27-95 mu- tant we isolated, but not the previously identified sec27-1 mutant. Previous EM studies on sec mutants with ER– Golgi apparatus trafficking defects have also shown that ER structure becomes disorganized at nonpermissive tem- perature (Novick et al., 1980; Duden et al., 1994). In con- trast to these mutants that affect traffic between the ER and Golgi complex, mutants with conditional defects in later steps of the secretory pathway (sec1-1, sec2-41, sec 3-2, sec4-8, sec6-4, sec 8-9, sec9-4, and sec15-1) did not show changes in ER morphology after three hours at non- permissive temperature (not shown). Besides its role in vesicular traffic from the Golgi appa- ratus to the ER (Lewis et al., 1997), Ufe1p has also been Prinz et al. Mutants Affecting ER Structure 467