(A) Fluorescence distribution of GFP-tagged endocytic cargoes (green) was analyzed in wildtype yeast cells expressing the vacuolar marker Vph1-mCherry (red). Cells were grown to mid-log phase at …
Top panels illustrate how PM fluorescence intensity was measured and calculated and bottom panels depict fluorescence intensity measurements over many cells (n > 30 cells).
Mup1 pHluorin signal originates almost exclusively from the PM (top panel).
Fluorescence distribution of GFP-tagged endocytic cargoes (green) was analyzed in wildtype yeast cells expressing the vacuolar marker Vph1-mCherry (red). Cells were grown to mid-log at 26°C and then …
For each heat-induced degradation timecourse experiment, the half-life of each cargo was estimated at each temperature using linear regression. Cargo half-life is shown plotted as a function of …
See also Figure 1—figure supplement 2.
(A) Fluorescence distribution of GFP-tagged endocytic cargoes (green) was analyzed in wildtype (left panels), rsp5-ww2 (middle panels), or rsp5-ww3 (right panels) yeast cells expressing the vacuolar …
(A) Yeast cells that were either grown at 26°C to mid-log phase (left), heated to 65°C for 10 min (middle), or treated with nystatin (right) were stained with propidium iodide (PI) and analyzed by …
(A) Heat-sensitivity analysis of wildtype and art mutant yeast cells. See Figure 4—figure supplement 1 for additional characterization of art mutant yeast cells. (B) Fluorescence distribution of …
Cells were grown to mid-log at 26°C and then shifted to 38°C for 2 hr. Plasma membrane (‘PM’) and Golgi localization are indicated.
Overexpression of ART2 can suppress the ts phenotype but not the canavanine hypersensitivity phenotype of Δart1 mutant cells.
(A) heat-sensitivity analysis of wildtype, Δart1, Δlyp1, and Δart1Δlyp1 mutant yeast cells. (B) Empty vector or plasmids encoding Lyp1-FLAG expressed from different promoters (pCPY, pLYP1, pADH1) …
(A) Model illustrating the major quality control mechanisms for integral membrane proteins: ERAD (proteasomal degradation), Golgi quality control (GQC; vacuolar/lysosomal degradation), and plasma …
Integral membrane proteins are subject to sequential quality control mechanisms including ERAD in the ER (I), GQC in the Golgi (II), and ART-Rsp5 mediated PMQC at the PM (III).
Cargo thermostability
Cargo | 34°C | 38°C | 40°C | 42°C |
Lyp1 | 160 | 50 | 25 | – |
Pma1 | Stable | 95 | 45 | – |
Mup1 | Stable | 100 | 65 | – |
Aqr1 | – | Stable | 100 | 70 |
Pdr5 | – | Stable | 400 | 200 |
Kinetic analysis of heat-induced cargo degradation (Figure 1—figure supplements 3 and 4) was used to estimate half-lives of each cargo at each temperature tested. Half-lives are indicated in minutes. ‘–’ indicates half-life not determined.