Fig. 4

Serotonin broadly inhibits proximity labelling of proteins, with a significant impact on SERT. (A) Representative images showing the effect of 10 µM serotonin alone and 10 µM serotonin with 500 µM Dz-PEG preincubation on biotinylation levels in HEK293T cells co-expressing HRP and SERT on the membrane. (B) Scatter plot analysis showing the correlation between NA and SERT intensity in (i) control, (ii) serotonin-treated, and (iii) Dz-PEG preincubation groups. Pearson correlation coefficient (r) and p-values are shown for each condition. The black line indicates a linear correlation. n = 19 (control), 15 (serotonin), 17 (serotonin + Dz-PEG) cells. (C) Heatmap showing the Log2 fold change of proteins enriched in control and Dz-PEG preincubation groups compared to the serotonin-treated group. Endogenously biotinylated proteins, including MCCC2, PCCB, MCCC1, PCCA, and PC, were used for normalization. Yellow indicates proteins with a Log2 fold change over 0.5. (D) Venn diagram showing the number of proteins with a Log2 fold change > 0.5 in control group (blue) and Dz-PEG preincubation group (green) compared to serotonin-treated alone group. (E) Quantification of normalised SERT abundance in control, serotonin-treated, and Dz-PEG preincubation groups. Scale bars represent 100 µm. n = 3 replicates in each condition from one experiment. Data are presented as mean ± SEM. One-way ANOVA with Tukey’s multiple comparisons test, *, **, *** represent p ≤ 0.05, 0.01 and 0.001 respectively.