Shiono M, et al. Supplementary Table S1

1. Shiono M, et al. Supplementary Table S1

2. Shiono M, et al. Supplementary Table S2

3. B C DAPI WT a b DAPI WT DAPI NSM DAPI NSM merge Tsc1 merge merge Tsc1 merge c D DAPI DAPI GSM GSM d DAPI merge Tsc1 merge Tsc1 f e DAPI NSM DAPI Y1571H Tsc1 merge Tsc1 merge Shiono M, et al. Supplementary Figure S1

4. y8 y7 y6 y5 y4 y3 y2 y1 1024 923 756 643 529 400 244 147 189 356 469 583 712 868 965 88 b1 b2 b3 b4 b5 b6 b7 b8 a [M+2H]2+ 84 80 75 70 S T pS L N E R P K 65 60 55 50 45 Intensity, counts 40 (y7-98)2+ 35 (b9-H2O)2+ (b6-NH3) 30 25 y72+ 20 y5 15 (a2-H2O) y2 y4 y6 y3 (y7-98) y7 10 5 0 100 200 300 400 500 600 700 800 900 m/z b y8 y7 y6 y5 y4 y3 y2 y1 [M+2H]2+ 894 763 596 539 482 345 288 175 12.0 11.0 10.0 88 219 386 443 500 637 694 807 9.0 b1 b2 b3 b4 b5 b6 b7 b8 8.0 7.0 (y9-98) Intensity, counts y2 (y7-98)2+ 6.0 y1 y3 5.0 (y7-98) y6 a4 S M pS G G H G L R 4.0 y5 (b8-H2O) 3.0 2.0 1.0 0.0 100 200 300 400 500 600 700 800 900 m/z Shiono M, et al. Supplementary Figure S2

5. a Rat genomic DNA Rat cDNA (exon 25, 31(-)) Rheb-GAP related domain 1 kb XhoI Not I BamHI Bgl II Bgl II BamHI BamHI BamHI BamHI ATG Probe TGA Tsc2 promoter SV40 poly-A signal G1556S Type : GSM b Founders obtained after GSM-Tg injection 23.1 Wild 9.4 Eker 6.6 4.4 Tg 2.3 2.0 (kb) c Wt GSM GSM (-) Wt (-) Tg : Tsc2 (~ 5.3 kb) Probe : Tsc2 SV40 poly-A Shiono M, et al. Supplementary Figure S3

6. a GSM-Tg : (-) (+) Tsc2 : +/Eker Eker/Eker Wt Eker b Tg : WT GSM IF: phospho- rpS6 (Ser235/ 236) c Tg : WT GSM IB: tuberin phospho- tuberin (Ser939) Shiono M, et al. Supplementary Figure S4