Reconstitution of the cargo recognition and packaging step of the Cvt pathway. (A) Localization of the cargo prApe1 in the MKO (atg15Δ RFP-APE1) strain, and localization of the receptor Atg19 and the adaptor Atg11 in the MKO (ape1Δ atg15Δ) strain. The MKO (atg15Δ RFP-APE1) strain and the MKO (ape1Δ atg15Δ) strain transformed with a plasmid expressing YFP-Atg19 (pYFPATG19(416)) or HA-tagged CFP-Atg11 (pCuHACFPATG11(414)) were grown in selective SMD medium to mid-log phase and observed by fluorescence microscopy. DIC, differential interference contrast. (B) Colocalization of prApe1, Atg19, and Atg11. The MKO (atg15Δ RFP-APE1) strain was transformed with a plasmid expressing YFP-Atg19, HA-tagged CFP-Atg11, or both, grown to mid-log phase and observed by fluorescence microscopy. When Atg19 was coexpressed with prApe1 in the MKO (atg15Δ RFP-APE1) strain, the cargo prApe1 colocalized with the receptor Atg19 (top). When Atg19 was absent, the adaptor Atg11 (arrows) did not colocalize with the cargo (middle). When prApe1, Atg19, and Atg11 were all present, the three proteins colocalized to the same structure (bottom). Bars, 2.5 μm.