COG4-Maid-mCherry (COG4-mAID) is functionally substituted endogenous COG4 and rapidly depleted upon IAA (auxin) treatment. (A) The diagram shows the development of the RPE1 COG4 KO cell line coexpressing COG4-mAID-mCherry (COG4-mAID) under COG4 promoter and OsTIR1–9myc. The COG4-mAid is ubiquitinated and degraded upon IAA treatment. (B) Expression of COG4-mAID rescues major cellular phenotypes associated with COG4 deficiency. WB shows the expression of COG4, myc, and COG-sensitive proteins in wild type, COG4 KO, and COG4-mAID cell lines. 10 μg of total cell lysates were loaded for each line. β actin has been used as a loading control. (C) COG4-mAID is Golgi localized (upper panel) and it is absent from the Golgi upon 1-h treatment with IAA. Airyscan superresolution IF analysis of COG4-mAID-mCherry (red) cells stained for GM130 (green). For better presentation, green and red channels are shown in inverted black and white mode whereas the merged view is shown in RGB mode. Scale bars, 20 μm. (D) WB of time-dependent depletion of COG4-mAID upon IAA treatment. A 10 μg of total cell lysates were loaded to each lane and probed with COG4 and actin antibodies. (E) The graph represents the quantification of D