Fig. 5: Ser754 phosphorylation modulates SCYL1 functions in Golgi structure maintenance.
From: mTORC1 controls Golgi architecture and vesicle secretion by phosphorylation of SCYL1
a Golgi structure depends on SCYL1 Ser754 phosphorylation. Two representative TEM pictures of MCF-7 SCYL1 KO cells re-expressing either HA-SCYL1(WT), HA-SCYL(S754A) or HA-SCYL1(S754E) upon 24 h doxycycline induction are shown. D, degradative compartment; E, endosome; G, Golgi apparatus; M, mitochondria; N, nucleus; PM, plasma membrane; #, lipid droplets. Scale bar: 500 nm. Additional pictures are available in Supplementary Fig. 5. b SCYL1 Ser754 phosphorylation maintains Golgi architecture. Contrary to HA-SCYL1(WT) and HA-SCYL1(S754E) proteins, which localize closer to the Golgi and around the nucleus, HA-SCYL(S754A) phospho-null localizes to the cell periphery. The enlarged Golgi observed in SCYL1 KO cells is rescued by HA-SCYL1(WT) and HA-SCYL1(S754E), but not by HA-SCYL1(S754A). Representative fluorescent micrograph of SCYL1 KO MCF-7 cells re-expressing HA-SCYL1(WT), HA-SCYL1(S754A) or HA-SCYL1(S754E) upon 24 h doxycycline induction. N = 3 biological replicates. c Quantification of b. The ratio between the Golgi volume, labeled with GOLGIN-97/GOLGA1 antibodies, and the total cellular volume, detected with the cell body detection tool (Imaris), was computed in individual cells. All results for SCYL1 KO MCF-7 cells re-expressing HA-SCYL1(WT), HA-SCYL1(S754A), or HA-SCYL1(S754E) for 24 h or 48 h were assembled in a violin plot (white dots within violins denote medians; top and bottom of violins represent the 25th and 75th percentiles; thick vertical lines within violins show the IQR, whereas thin vertical lines represent the rest of the distribution extending the 1.5xIQR). Single-cell values are indicated with small white dots. An unpaired two-tailed Student’s t-test was used for comparison: 24 h + WT/VC: p = 0.00037(***) and /S754A: p = 0.0036(**); 48 h + WT/VC: p = 9.72e-14(****) and /S754A: p = 2.98e-06(****); 48 h + S754A/VC: p = 0.00066(***); 48 h + S754E/VC: p = 2.21e-29(****) and /S754A: p = 5.37e-17(****). ≥30 cells were measured amongst technical triplicates of two biological replicates (n = 6). Error bars = SEM. Source data are provided as Source Data file.
