Figure S1.
VAMP2 is not involved in delivery of receptors from the biosynthetic pathway. For Fig. 2. (A) VAMP2 enrichment (left) in TfR puffs in cells co-expressing TfR-SpH and VAMP2-pHuji co-expressing cells pretreated with 10 µM CHX for 2 h before imaging (+CHX, median ± 95% CI, puff = 67, 2 cells from one experiment). Proportion of puffs > or ≤ cutoff = 1× SDbaseline is shown in the pie chart (right). (B) Proportion of puffs from TfR-SpH and VAMP2-pHuji co-expressing cells non-treated with CHX (−CHX). Proportion > cutoff = 1× SDbaseline is almost identical for cells treated or non-treated with CHX. (C) Representative Western blot probing a fast turnover protein GRK3 with GAPDH as internal control in whole-cell lysates prepared from cells either treated with vehicle (EtOH) for 2 h or 10 µM CHX for indicated time. (D) Quantification of relative GRK3 levels by first normalizing to GAPDH levels then to vehicle control (±SD, n = 3 independent experiments). (E and F) Representative confocal slices of PC12 cells immunostained for endogenous VAMP2 and cis-Golgi marker GM130 (E) or tran-Golgi network marker TGN38 (F). Scale bar = 10 µm. Denoted sections (yellow squares and enlarged) were analyzed by pixel-by-pixel fluorescence correlation between the VAMP2 channel and the Golgi marker channels. Pixel fluorescence intensity was normalized to the maximum fluorescence and plotted in scatter plots on the right. The dashed diagonal line indicates perfect colocalization, and VAMP2 shows very little to no colocalization with GM130 or TGN38. Source data are available for this figure: SourceData FS1.

VAMP2 is not involved in delivery of receptors from the biosynthetic pathway. For Fig. 2. (A) VAMP2 enrichment (left) in TfR puffs in cells co-expressing TfR-SpH and VAMP2-pHuji co-expressing cells pretreated with 10 µM CHX for 2 h before imaging (+CHX, median ± 95% CI, puff = 67, 2 cells from one experiment). Proportion of puffs > or ≤ cutoff = 1× SDbaseline is shown in the pie chart (right). (B) Proportion of puffs from TfR-SpH and VAMP2-pHuji co-expressing cells non-treated with CHX (−CHX). Proportion > cutoff = 1× SDbaseline is almost identical for cells treated or non-treated with CHX. (C) Representative Western blot probing a fast turnover protein GRK3 with GAPDH as internal control in whole-cell lysates prepared from cells either treated with vehicle (EtOH) for 2 h or 10 µM CHX for indicated time. (D) Quantification of relative GRK3 levels by first normalizing to GAPDH levels then to vehicle control (±SD, n = 3 independent experiments). (E and F) Representative confocal slices of PC12 cells immunostained for endogenous VAMP2 and cis-Golgi marker GM130 (E) or tran-Golgi network marker TGN38 (F). Scale bar = 10 µm. Denoted sections (yellow squares and enlarged) were analyzed by pixel-by-pixel fluorescence correlation between the VAMP2 channel and the Golgi marker channels. Pixel fluorescence intensity was normalized to the maximum fluorescence and plotted in scatter plots on the right. The dashed diagonal line indicates perfect colocalization, and VAMP2 shows very little to no colocalization with GM130 or TGN38. Source data are available for this figure: SourceData FS1.

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