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Supplementary Information to
Mapping the interaction of Snf1 with TORC1 in Saccharomyces
cerevisiae
Jie Zhang1, Stefania Vaga2, Pramote Chumnanpuen1, Rahul Kumar1, Goutham N. Vemuri1, Ruedi
Aebersold2, 3, Jens Nielsen1, *
1
Department of Chemical and Biological Engineering, Chalmers University of Technology, Kemivägen
10, SE-412 96, Göteborg, Sweden
2
Department of Biology, Institute of Molecular Systems Biology, ETH Zürich, 8093 Zürich,
Switzerland
3
Faculty of Science, University of Zürich, Switzerland
*Corresponding author. Department of Chemical and Biological Engineering, Chalmers University of
Technology, Kemivägen 10, SE-412 96, Gothenburg, Sweden. Tel.: +46 31 772 38 04; Fax: +46 31 772
38 01; E-mail: [email protected]
1
Supplementary Figures
Supplementary figure S1 The mutant strain tor1∆ is more sensitive to rapamycin treatment. The reference
(A)and tor1Δ (B) strains were grown in media with different concentrations of rapamycin (only results of up to
5 nM of rapamycin treatment were shown here).
2
Supplementary figure S2 Distribution of variance in -omics data among each principal component. Principal
component analysis (PCA) was applied to decrease the dimension of the data set and therefore to decrease
the redundancy of the (A) transcriptome data; (B) phosphoproteome data. Only the first 10 principal
components are plotted.
Supplementary figure S3 Expression of key genes involved in acetate metabolism from all strains and both Cand N-limited conditions. The numbers represent relative expression levels (log 2 fold change) in the mutant
strains with respect to the reference strain, where blue indicates the gene lower expressed while red being
higher expressed in the mutant strains.
3
Supplementary figure S4 Phosphorylation of proteins involved in regulation of gene transcription. Numbers
indicate the significance of change in phosphorylation level, where yellow (positive values) indicates higher
and green (negative values) indicates lower phosphorylation in mutant strain with respect to the reference
strain.
4
Supplementary figure S5 Phosphorylation of proteins that interact with Snf1, TORC1 and PKA in mutant
strains grown on C-limited condition. Numbers indicate the significance of change in phosphorylation level,
where yellow (positive values) indicates higher and green (negative values) indicates lower phosphorylation in
mutant strain with respect to the reference strain.
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Supplementary figure S6 Expression of amino acid biosynthetic gene. Numbers are average values from the
biological replicates for all strains on both C- and N-limited conditions. Red (positive values) indicates higher
expression while blue (negative values) indicates lower expression in mutant strain with respect to the
reference strain.
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Supplementary figure S7 Phosphorylation of proteins involved in different processes. (A) Proteins in lysine
synthesis; (B) Proteins involved in fatty acid synthesis and β-oxidation. The color indicates the relative
abundance of the phosphorylated peptides, yellow being higher and black being lower phosphorylated.
7
Supplementary Tables
Supplementary Table S1 Genotypes of the strains used in this study.
Strain
CEN.PK 113-7D
CEN.PK 506-1C
CEN.PK 113-1A
BY4741
CEN.PK JZH-F1
CEN.PK JZH-F2
CEN.PK JZH-G1
Genotype
MATa URA3 HIS3 LEU2 MET15
MATa URA3 HIS3 LEU2 MET15; snf1Δ::KanMX
MATα URA3 HIS3 LEU2 MET15
MATa ura3 his3 leu2 met15; tor1Δ::KanMX
MATa URA3 HIS3 LEU2 MET15; tor1Δ::KanMX
MATα URA3 HIS3 LEU2 MET15; tor1Δ::KanMX
MATa URA3 HIS3 LEU2 MET15; snf1Δtor1Δ::KanMX
8
Source
Van Dijken et al
Usaite et al
Matthias Rose
Matthias Rose
This study
This study
This study
Supplementary table S2 Reporter metabolites and effectors in mutant strains compared to the reference. All identified reporter X had significance count larger than 1 to
reduce the bias from the size of the sub-network. Cutoff is set to be Z-score > 1.5 (metabolite), p-value < 0.05 (TF) and p-value < 0.01 (biological process). The subscript
indicates the location of the metabolite, e.g., ‘xt’: extracellular; ‘M’: mitochondrial. [acp]: acyl-carrier protein; αKG: α-ketoglutarate; GSH: glutathione; CDK: cyclindependent kinase
Reporter X (significance count ≥ 2)
Metabolite (Z-score > 1.5)
C-lim
N-lim
+
+
snf1∆
NADP , NADPH, NAD , NADH, CoA, αKG, L-Asp, LGln, CMP, L-Asn, Oxaloacetate, L-Ornithine,
Fecosterol
snf1∆
tor1∆
CoA, αKG, Gluxt, 1L-myo-Inositol 1-P, NADP+,
NADPH, Alaxt, L-Glu, L-Thr
snf1∆
Dolichyl-P, GSH, Xanthosine 5'-P, Phosphatidate, 4Aminobutanoate, Oxidized GSH, Malate, Alaxt,
Dolichyl β-D-mannosyl-P, IsocitrateM, αKGM,
Mannan, MalateM, GABAxt, L-Gln, Decanoyl-[acp],
Dodecanoyl-[acp], Myristoyl-[acp], Hexadecanoyl[acp], Palmitoyl-[acp], Stearoyl-[acp], Oleoyl-[acp],
Linolenoyl-[acp], (R)-LactateM, Asnxt, Glnxt, CitrateM,
H2O2, Urea, L-Ser, CoAM, Ferricytochrome CM,
Ferrocytochrome CM, N-(L-Arginino)succinate
αKG, MalateM, Malate, Acetaldehyde, Acetyl-CoA
tor1∆
TF (p-value < 0.05)
Biological process (p-value < 0.01)
Msn2/4, Pho4, Ino2/4, Hap1, Gcn5, Cat8,
Rtg1/3, Pho90, Opi1, Cdc39, Ras2, Haa1,
Mks1, Taf12, Oaf1, Snf7, Pho87, Pho91, Rpt6,
Mot2, Fus3
Msn2/4, Oaf1, Pip2, Hap1, Cat8, Rtg1-3, Rnt1,
Cdc39, Pho4, Ssy1, Ino2/4, Rsf1, Taf12, Hap23, Mot3, Hog1, Fus3, Gts1, Sok2, Ama1, Yap1,
Mot2, Spt6, Pho90, Mks1, Plc1, Ras2, Ssn3
Hap2-5, Pho90, Pbs2, Msn2/4, Rnt1, Swi1,
Pho87, Pho91, Ssa1, Pho4, Grr1, Fks1, Ras2,
Rtg1-2, Rgt1, Ptr3, Plc1, Pkc1, Yap1, Mks1,
Rlm1, Ino4, Snf5, Ire1, Uga3, Alpha1, Cin5,
Std1, Snf6, Crz1, Cup9, Snf2, Cat8, Ssn3, Dal81
Glu biosynthesis, ergosterol biosynthesis, response to stress, aerobic respiration,
glucose metabolism, cellular response to nitrogen starvation
Gln3, Dal80, Hst1, Pgd1, Gal11, Aro80, Snf1/4,
Med2, Rcs1, Cse2, Ppz1, Ppz2, Spt20, Sin4,
Snf2, Pdr1
cytokinesis, completion of separation, cell-substrate adherens junction assembly,
high-affinity iron ion transport, mitosis, TCA cycle, nicotinamide riboside metabolism,
Arg biosynthesis, Glu biosynthesis, pyridoxine metabolism, cell morphogenesis
during conjugation, ethanol metabolism, His biosynthesis, isocitrate metabolism,
ammonium transport, regulation of G-protein coupled receptor protein signaling
pathway, nitrogen compound metabolism, NAD biosynthesis, thiamin biosynthesis,
ribosomal large subunit assembly and maintenance, rRNA processing, L-Phe
catabolism, allantoin catabolism, sulfate assimilation, regulation of CDK activity
9
aerobic respiration, nitrogen utilization, Glu biosynthesis, ammonium transport,
ergosterol biosynthesis, response to drug, ethanol metabolism, pheromonedependent signal transduction during conjugation with cellular fusion, glucose
metabolism
TCA cycle, ATP synthesis coupled proton transport, glucose metabolism, Glu
biosynthesis, response to oxidative stress, carbohydrate metabolism,
posttranslational protein targeting to membrane, GPI anchor biosynthesis, aerobic
respiration, phosphate transport, cell cycle arrest, protein amino acid glycosylation,
response to unfolded protein, αKG metabolism, rRNA processing, protein targeting
to ER, acetaldehyde metabolism
snf1∆
tor1∆
αKGM, L-Asn, 4-Aminobutanoate, Alaxt, L-Ala, Hisxt,
GSH, Glyxt, GABAxt, CitrateM, IsocitrateM, L-Gln,
Malate, Dolichyl-P, D-Glucono-1,5-lactone 6-P,
Asnxt, Glnxt, Xanthosine 5'-P, 6-P-D-gluconate, LCitrulline, Oxidized GSH, MalateM, Thrxt, Dolichyl βD-mannosyl-P, NADHM, N-(L-Arginino)succinate,
OrthophosphateM, Phosphatidate, Oxaloglutarate,
NH3xt, Urea, L-Pro, Mannan, OxaloacetateM, (R)LactateM, Proxt, Glycogen, CDP choline
Msn2/4, Arg82, Ure2, Pho90, Gln3, Rtg1-3,
Gat1, Plc1, Rnt1, Hog1, Grr1, Hap2-5, Mac1,
Ssa1, Dal80, Swi1, Cup9, Pbs2, Arg80, Arg81,
Gzf3, Gas1, Ptr3, Mks1, Yap1, Pho87, Pho91,
Snf6, Rlm1, Kcs1, Fks1, Std1, Vps24, Rgt1,
Pho4, Ire1, Pkc1, Rgt2, Uga3, Cin5, Alpha1,
Ppz1, Ppz2, Snf2, Snf5, Rds3, Pdr1
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TCA cycle, ATP synthesis coupled proton transport, aerobic respiration, glucose
metabolism, cellular response to nitrogen starvation, carbohydrate metabolism, GPI
anchor biosynthetic process, SRP-dependent cotranslational protein targeting to
membrane translocation, copper ion import, rRNA processing, Glu biosynthesis,
nitrogen compound metabolism, fatty acid transport, pentose-phosphate shunt,
oxidative branch, rRNA processing, response to unfolded protein, response to
oxidative stress, citrate metabolism
1
2
Supplementary table S3 Concentration of free intracellular amino acids for all strains grown on both C- and N-limited conditions. The numbers are average values of two
technical replicates (μmole gCDW-1).
reference
AA
C-lim
snf1Δ
tor1Δ
Rep_1
Rep_2
Rep_3
Rep_1
Rep 2
Rep 3
Rep_1
Rep_2
Rep_3
Rep_1
Rep 2
Rep 3
Alanine
20.04921
19.94934
21.16735
4.480694
4.478226
5.035942
19.47548
13.0102
11.66243
6.408503
6.443236
5.203967
Glycine
3.641368
3.708185
3.667481
2.278083
1.908752
2.691387
4.048956
5.341926
5.475709
2.720675
2.486874
2.731341
Valine
6.761814
7.55951
7.265117
2.430721
2.443289
2.612772
7.493584
6.023365
5.740408
2.521538
2.337062
2.729697
Leucine
0.832947
0.991025
0.882971
0.437717
0.417314
0.459556
1.05511
1.123639
1.231968
0.578937
0.435109
0.489828
Isoleucine
1.722463
1.957116
1.745663
0.658676
0.574814
0.709591
2.442844
1.864013
2.819015
0.699062
0.729957
0.75798
Threonine
3.361731
3.846086
3.766883
2.199802
2.542463
2.847987
4.346092
5.562616
5.285922
2.83932
2.856912
2.904568
Proline
4.13944
4.795164
4.849855
0.430723
0.356386
0.487981
3.971932
2.979007
3.155479
0.620766
0.512538
0.861495
Asparagine
4.905676
4.935045
4.961107
0.118294
<0.001
<0.001
4.593949
4.486555
3.283613
<0.001
<0.001
<0.001
Methionine
0.686888
1.378075
0.692512
0.603617
0.60735
0.835208
0.718093
1.552101
0.624643
0.726927
0.732301
0.800085
Aspartic Acid
15.80237
17.92585
17.85312
7.378818
9.023323
9.048099
18.32422
13.68614
14.99866
9.410176
11.0426
8.938164
Phenylalanine
1.494339
1.485036
1.559518
1.678714
1.58714
1.935133
1.513703
1.42322
1.476826
1.881517
1.779256
1.807002
Glutamic acid
148.6233
179.8395
171.2055
35.17783
44.08228
42.94568
142.4655
114.1701
117.2037
37.44567
40.80106
40.96007
Glutamine
49.67352
106.5039
80.82594
2.129589
5.459775
6.375537
37.43623
40.77015
41.71903
4.079312
9.123572
5.755484
Lysine
2.870628
3.289582
3.958193
1.508322
1.577135
1.662368
3.150539
4.957236
4.516123
2.33041
2.230124
1.965724
Histidine
5.479834
12.13147
11.83719
1.55072
1.330378
1.659969
3.796214
3.713023
3.682798
1.563664
1.632059
1.398922
Tyrosine
1.636134
1.836987
1.900938
0.658678
0.640488
0.750447
1.625089
1.2154
1.423272
0.664601
0.637394
0.684394
Tryptophan
0.190292
0.352524
0.304484
0.001782
0.016121
<0.001
0.136562
0.127733
0.126834
<0.001
0.005578
<0.001
3
4
snf1Δtor1Δ
To be continued on the next page.
5
11
reference
AA
N-lim
snf1Δ
tor1Δ
snf1Δtor1Δ
Rep_1
Rep_2
Rep_3
Rep_1
Rep_1
Rep_2
Rep_3
Rep_1
Rep_1
Rep_2
Rep_3
Rep_1
Alanine
24.08213
26.85776
25.59549
11.93622
10.15968
12.78845
15.97484
16.50682
18.99548
9.81626
10.67418
9.847464
Glycine
4.783765
4.405195
4.652467
3.908729
3.596203
4.441132
6.322932
5.885758
6.452478
2.890736
3.079233
3.272792
Valine
9.65326
8.706566
9.151242
4.444143
4.697154
5.431461
5.77775
5.842978
7.340943
3.225069
3.780902
3.703619
Leucine
1.384566
1.334291
1.272173
0.712171
0.775175
0.920386
1.554827
1.285662
1.606889
0.674515
0.619579
0.727416
Isoleucine
2.555792
2.404078
2.457168
0.646916
0.951925
1.063829
4.63701
2.807102
3.026147
0.852795
0.967477
1.173769
Threonine
4.397919
4.473898
4.665797
2.469272
5.295307
5.91696
3.792697
5.196004
6.275091
3.864195
4.356017
4.349844
Proline
6.008796
5.76896
6.496845
1.529604
1.332278
1.679015
2.011619
3.431462
4.669693
0.931081
1.345634
1.164968
Asparagine
6.685512
7.71806
6.952898
<0.001
0.039433
0.111891
5.218463
5.378189
6.050602
<0.001
<0.001
<0.001
Methionine
0.860852
0.833431
1.426569
0.736392
0.620212
0.660811
0.952287
1.430555
1.441611
0.687316
0.587913
0.667432
Aspartic Acid
17.84565
11.53562
18.70147
17.16092
17.07096
19.51797
11.94498
11.0406
16.7356
14.08627
18.26311
20.22585
Phenylalanine
1.923499
1.659729
1.784313
1.710829
1.546645
1.869282
0.604854
1.351235
1.302443
1.786341
1.649587
0.891169
Glutamic acid
157.1198
169.98
176.916
74.85193
71.49478
78.79179
124.5101
121.6797
166.279
50.26728
58.19312
77.00248
Glutamine
79.89603
113.4605
146.4862
49.23778
55.04806
68.24478
43.91489
73.36365
113.5709
18.57646
25.7291
41.26609
Lysine
5.997851
6.198438
5.728678
5.51852
5.924457
7.563857
4.07708
4.699813
5.083411
2.80198
1.890494
2.953335
Histidine
11.51288
9.763431
20.9897
3.702733
2.82053
6.883333
4.056512
2.4549
5.421118
2.554758
0.823334
2.385934
Tyrosine
2.447718
2.120823
2.30658
0.657784
0.704561
0.895781
1.183455
1.519277
1.332838
0.749121
0.671018
0.770529
Tryptophan
0.319529
0.328617
0.412486
0.019423
0.023317
0.025205
0.093682
0.090812
0.09494
<0.001
0.025575
0.013866
1
2
3
12
1
2
Supplementary table S4 Concentration of fatty acids for all strains grown on both C- and N-limited conditions. The numbers are average values of two technical
replicates (μmole gCDW-1).
reference
C-lim
snf1Δ
3
snf1Δtor1Δ
Rep_1
Rep_2
Rep_3
Rep_1
Rep_1
Rep_2
Rep_1
Rep_2
Rep_3
Rep_1
Rep_1
Rep_2
C14:0
0.229116
0.119922
0.122056
0.081277
0.099083
0.110378
0.222777
0.209388
0.243512
0.128919
0.083412
0.035803
C16:0
2.255683
1.654994
2.177783
2.044178
1.638204
2.924643
2.005399
1.637855
1.773149
5.590092
3.522292
2.005572
C16:1*
1.989374
1.203035
1.517198
1.471519
2.762099
2.390537
3.081363
0.935239
1.520259
0.601744
2.60672
0.562301
C18:0
0.47364
0.705511
0.656909
1.234149
3.107187
2.786736
0.457345
0.380259
0.406573
3.901257
3.267356
4.487907
C18:1
7.970918
4.450917
5.2346
23.03356
74.9425
62.45083
8.910085
3.777247
4.430847
11.72585
63.22927
14.49676
C20:0
0.004512
<0.001
0.001504
0.013967
0.031973
0.054525
0.001518
0.001772
0.001711
<0.001
0.0297
0.045165
C22:0
0.004487
<0.001
<0.001
0.003217
0.002221
0.019894
<0.001
0.001396
<0.001
<0.001
0.014089
0.027679
FA
Rep_1
Rep_2
Rep_3
Rep_1
Rep_1
Rep_2
Rep_3
Rep_1
Rep_1
Rep_2
Rep_3
Rep_3
C14:0
0.102296
0.254648
0.235496
0.073707
0.024367
0.109963
0.062053
0.248201
0.177788
0.102637
0.07549
0.020944
reference
N-lim
tor1Δ
FA
snf1Δ
tor1Δ
snf1Δtor1Δ
C16:0
3.83245
3.543685
4.280376
3.391195
2.269874
3.429316
3.664327
4.918738
3.157535
3.498184
3.948075
4.367951
C16:1*
1.658994
1.617028
3.20578
1.264531
0.44646
1.665703
0.575
2.659068
1.864091
1.439356
1.455165
2.159919
C18:0
1.600061
1.540023
1.639863
2.411334
4.552516
2.583977
1.705149
1.776812
1.032734
3.208988
5.148261
6.101613
C18:1
19.79423
14.79566
21.89869
38.32071
11.11959
27.94223
7.849762
23.24861
0.728749
67.36435
66.03698
85.48892
C20:0
<0.001
0.005014
<0.001
0.024526
0.040896
0.035887
0.006928
0.004872
0.004247
0.037008
0.024963
0.012989
C22:0
<0.001
<0.001
<0.001
0.010441
0.004242
0.008154
<0.001
0.001536
0.002276
0.018005
0.016528
0.000523
* All measurements are absolute values except that C16:1 was only relatively quantified.
13