Fig. 5
ATG9A and SERINC colocalisation in peripheral puncta is dependent on AP-4. CRISPR/Cas9 gene editing was used to introduce a C-terminal Clover (modified GFP) tag to endogenous SERINC1 or SERINC3 in HeLa cells. Cells were transfected with siRNA to knock down AP-4, or were mock transfected (without siRNA) as a control. a Confocal microscopy with Airyscanning was used to image SERINC1-Clover (via anti-GFP) and anti-ATG9A. Representative images show a confocal slice of the whole cell, and a peripheral 10 × 10 μm square imaged with Airyscanning in Superresolution mode, used for the quantification of colocalisation between SERINC1-Clover and ATG9A. Scale bar: 10 μm. b Quantification of colocalisation between SERINC1-Clover and ATG9A in peripheral regions of mock treated and AP-4 knockdown (KD) cells, using Thresholded Pearson’s Correlation Coefficient. 20 cells were quantified per condition. Data show mean (n = 20), and results of a two-tailed Mann–Whitney U-test: ***p ≤ 0.001. c Confocal microscopy with Airyscanning was used to image SERINC3-Clover (via anti-GFP) and anti-ATG9A, as in a. d Quantification of colocalisation between SERINC3-Clover and ATG9A in peripheral regions of mock treated and AP-4 knockdown cells, using Thresholded Pearson’s correlation coefficient. 19 cells were quantified per condition. Data show mean (n = 19), and results of a two-tailed Mann–Whitney U-test: ***p ≤ 0.001
