Cdc1 removes the ethanolamine phosphate of the first mannose of GPI anchors and thereby facilitates the integration of GPI proteins into the yeast cell wall.
Bottom Line: Cdc1-314(ts) mutants do not accumulate GPI proteins in the ER but have a partial secretion block later in the secretory pathway.Osmotic support restores GPI protein secretion and actin polarization but not growth.This suggests that the presumed transglycosidases Dfg5 and Dcw1 of cdc1-314(ts) transfer GPI proteins to cell wall β1,6-glucans inefficiently.
Affiliation: Department of Biology, University of Fribourg, CH-1700 Fribourg, Switzerland.Show MeSH
Mentions: Because ERAD mutants rescued growth of cdc1 (Figure 3F), we hypothesized that ERAD mutants may stabilize Cdc1-314. To verify this, we expressed HA-Cdc1 or HA-Cdc1-314, that is, tagged versions carrying the hemagglutinin-derived HA epitope (HA) under the native promoter from a centromeric vector in WT cells and quantified the amounts of HA-tagged protein after shifting cells from 24° to 30° or 37°C, as shown in Figure 4A. In cells growing at 24°C, Cdc1-314 already was 3.8-fold less abundant than WT Cdc1. A shift to 30°C did not destabilize Cdc1-314, although cells depending only on mutant protein do not grow at this temperature (Figure 3A). When cells were shifted to 37°C, a third of Cdc1-314 was degraded within 15–30 min, but the protein remained stable thereafter. The same temperature shifts reduced the amounts of WT Cdc1, but more slowly, so that it took more than 4 h to reach equilibrium levels.
Affiliation: Department of Biology, University of Fribourg, CH-1700 Fribourg, Switzerland.