A Blot#2
2D
3D
2D
3D
2D
3D
2D
3D
2D
3D
2D
3D
ERK1/2
rpS6
αTubulin
p70S6K
4E-BP1
p-AktS473l.e.
Akt
p-PRAS40T246l.e.
αTubulin
E-Cadherinl.e.p-ERK1/2T202/Y204s.e.p-ERK1/2T202/Y204l.e.
p-AktT308
p-p70S6KT389l.e.
p-rpS6S240/244
p-p70S6KT389s.e.
E-Cadherins.e.
p-PRAS40T246s.e.
p-AktS473s.e.
2D
3D
2D
3D
2D
3D
2D
3D
2D
3D
2D
3D
Blot#3
ERK1/2
rpS6
αTubulin
p70S6K
4E-BP1
Akt
p-PRAS40T246l.e.
αTubulin
E-Cadherinl.e.
p-ERK1/2T202/Y204s.e.p-ERK1/2T202/Y204l.e.
p-AktT308
p-p70S6KT389l.e.
p-rpS6S240/244l.e.
p-p70S6KT389s.e.
E-Cadherins.e.
p-PRAS40T246s.e.
p-AktS473s.e.
GAPDH
GAPDH
p-rpS6S240/244s.e.
p-PKC(pan)βIIS660
B
Suppor(ngFigureSF1(figshare)–AnalysisofPI3K/AKT/mTOR-andRAS/RAF/MAPK-singalingin2Dvs3D cell culture. The colon cancer cells (LS174T,HT29, SW620,HCT116,Caco2andDLD-1)werecultured in 2D and 3D for 48h. Cells and spheroids were harvested andWestern blot analysis ofdifferentproteinsof thePI3K/AKT/mTOR-andRAS/RAF/MAPK-pathwaywascarriedout.Detec_onof phosphorylated proteins is indicated by p- prior to the protein name and the phosphorylatedaminoacid is indicatedbehind theproteinname.α-tubulin andGAPDHservedas loading control.Twoindependentbiologicalreplicatesareshown(A,B).
