Fig. 4.

Transport of Cps1 to the vacuole is impaired in sur4Δ vps21Δ and fen1Δ vps21Δ mutants. (A) BY4741 (wild-type), 5763 (fen1Δ), 5281 (sur4Δ), 1865 (vps21Δ), MRY90 (fen1Δ vps21Δ), and MRY91 (sur4Δ vps21Δ) cells, each harboring pOK489 (mRFP-Cps1), were grown at 30°C to log phase and subjected to fluorescence microscopy. Left panels, DIC images; right panels, mRFP fluorescence. Bar, 5 μm. (B) Total lysates were prepared from cells indicated in (A), separated by SDS-PAGE, and detected by immunoblotting with anti-DsRed antibody or, to demonstrate uniform protein loading, anti-Pgk1 antibody. Expression levels of mRFP-Cps1 were significantly low in the fen1Δ and sur4Δ mutants, compared with the other strains used, for an unknown reason. Thus, the amounts of lysates subjected to SDS-PAGE were increased for the fen1Δ and sur4Δ mutants (3- and 4-fold for the fen1Δ and sur4Δ cells, respectively) to make the total amount of mRFP-Cps1 plus mRFP comparable to the other strains.