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. 2020 Jun 8;295(30):10406–10419. doi: 10.1074/jbc.RA120.012575

Figure 2.

Figure 2.

Sec61, Sec66, and Sec72 are required for the translocation of N-terminal signal peptide-deprived proteins into the ER. A, Western blot analysis of CPY*ΔN28 expressed in the cold-sensitive sec61-41 mutant. CPY*ΔN28 was expressed in the indicated cells at the permissive temperature (30 °C). Extracts from these cells were resolved by SDS-PAGE, and CPY*ΔN28 was visualized by immunoblotting using an anti-V5 antibody. B, Western blot analysis of CPY* expressed in the sec61-41 mutant. CPY* was expressed and analyzed as described above. C, Western blot analysis of Mid1ΔN23 expressed in the sec61-41 mutant. Mid1ΔN23-HA was expressed and analyzed as described above. D, Western blot analysis of WT Mid1 expressed in the sec61-41 mutant. Mid1-HA was expressed and analyzed as described above. E, Western blot analysis of CPY*ΔN28 expressed in the sec66 and sec72 mutants. CPY*ΔN28-V5 was expressed and analyzed as described above. F, Western blot analysis of CPY* expressed in the sec66 and sec72 mutants. CPY*-V5 was expressed and analyzed as described above. G, Western blot analysis of Mid1ΔN23 expressed in the sec66 and sec72 mutants. Mid1ΔN23-HA was expressed and analyzed as described above. H, Western blot analysis of WT Mid1 expressed in the sec66 and sec72 mutants. Mid1-HA was expressed and analyzed as described above. I, Western blot analysis of CPY*ΔN28 in the get and snd mutants. CPY*ΔN28-V5 was expressed in the get and snd mutants. Extracts from the mutants were analyzed as described above. The ratio of glycosylated versus total CPY*ΔN28 proteins was represented as a bar graph. n represents the number of biological replicates.