Role of Erv29p in Collecting Soluble Secretory Proteins into ER-Derived Transport Vesicles William J. Belden and Charles Barlowe

Supplementary Material

Supplemental Figure 1. Cells lacking Erv29p accumulate ER forms of gpf and CPY but not Gas1p or invertase. Cell extracts were prepared from FY834 (WT), CBY966 (erv29), and RSY263 (sec12-4) after a 10-min pulse with ³⁵S-labeled amino acids and chased for indicated times (8). (A) gpf, (B) CPY, and (C) Gas1p were immunoprecipitated from a common extract and resolved on 10% polyacrylamide gels. (D) Invertase pulse-chase analyses were performed with derepressed cells. Proteins were visualized and transport rates quantified using a PhosphorImager. Data plots of these experiments are included in the supplemental material (20).

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Supplemental Figure 2. Expression level of Erv29p influences the transport kinetics of gpf. (A) Cells were pulsed for 5 min with ³⁵S-labeled amino acids then chased for indicated times. Proteins were immunoprecipitated from extracts prepared from FY834 (WT), CBY966 (erv29), and CBY1161 (WT, MF1-2), a strain that overproduces gpf. (B) As in (A), with CBY1162 (MF1-2, ERV29 × 2), a strain that overproduces gpf and contains two chromosomal copies of ERV29, included. Proteins were visualized and transport rates quantified using a PhosphorImager. Data plots of these experiments are included in the supplemental material (20).

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