Background
Methods
Melanoma samples
Cell lines
Stable transduction of cell lines
RNA extraction, amplification, labelling and hybridisation
Microarray analysis
Western blot analysis
Real-time PCR
Cell proliferation assays
Results
Transcript expression of P53 target genes in metastatic melanoma
No. | Accession No. | Gene Symbol | Gene Name | Fold change | p-value |
---|---|---|---|---|---|
Apoptosis
| |||||
1
| NM_001040619.1 | ATF3 | Activating transcription factor 3, transcript variant 4 | 4.62 | 0.0042 |
2
| NM_003879.3 | CFLAR/FLIP | CASP8 and FADD-like apoptosis regulator | 3.10 | 0.0063 |
3
| NM_004324.3 | BAX | BCL2-associated X protein, transcript variant beta | -3.92 | 0.0127 |
4
| NM_138578.1 | BCL2L1 | BCL2-like 1 (Bcl-xL), transcript variant 1 | -5.77 | 0.0144 |
5
| NM_001226.3 | CASP6 | Caspase 6, transcript variant alpha | -13.07 | 4.98E-05 |
6
| NM_033340.2 | CASP7 | Caspase 7, transcript variant beta | -3.69 | 0.0465 |
7
| NM_001080125.1 | CASP8 | Caspase 8, transcript variant G | -11.38 | 2.28E-05 |
8
| NM_033356.3 | CASP8 | Caspase 8, transcript variant C | -2.96 | 0.0273 |
9
| NM_001007277.1 | EI24 | Etoposide induced 2.4 mRNA, transcript variant 2 | -2.81 | 0.0245 |
10
| NM_021127.1 | PMAIP1 | Phorbol-12-myristate-13-acetate-induced protein 1 (Noxa) | 12.18 | 0.0063 |
11
| NM_000314.4 | PTEN | Phosphatase and tensin homolog | -3.29 | 0.0062 |
12
| NM_003840.3 | TNFRSF10D | Tumor necrosis factor receptor superfamily, member 10d | -29.89 | 8.37E-07 |
13
| NM_004881.2 | TP53I3 | Tumor protein p53 inducible protein 3, transcript variant 1 | -4.50 | 0.0097 |
Cell cycle, proliferation and differentiation
| |||||
14
| NM_033031.2 | CCNB3 | Cyclin B3, transcript variant 3 | -8.38 | 1.47E-04 |
15
| NM_033031.2 | CCNB3 | Cyclin B3, transcript variant 3 | -10.79 | 1.18E-04 |
16
| NM_001759.2 | CCND2 | Cyclin D2 | 76.10 | 1.73E-07 |
17
| NM_001238.1 | CCNE1 | Cyclin E1, transcript variant 1 | -7.61 | 2.07E-10 |
18
| NM_057749.1 | CCNE2 | Cyclin E2 | 3.54 | 0.0476 |
19
| NM_199246.1 | CCNG1 | Cyclin G1, transcript variant 2 | -5.75 | 0.0146 |
20
| NM_004354.1 | CCNG2 | Cyclin G2 | -10.66 | 2.10E-04 |
21
| NM_001798.2 | CDK2 | Cyclin-dependent kinase 2, transcript variant 1 | -4.09 | 0.0476 |
22
| NM_001798.2 | CDK2 | Cyclin-dependent kinase 2, transcript variant 1 | -17.99 | 0.0017 |
23
| NM_058197.3 | CDKN2A | Cyclin-dependent kinase inhibitor 2A, transcript variant 3 | 4.97 | 0.0202 |
24
| NM_058195.2 | CDKN2A | Cyclin-dependent kinase inhibitor 2A, transcript variant 4 | -7.09 | 0.0351 |
25
| NM_015675.2 | GADD45B | Growth arrest and DNA-damage-inducible, beta | 7.41 | 0.0020 |
26
| NM_006705.2 | GADD45G | Growth arrest and DNA-damage-inducible, gamma | 11.13 | 0.0154 |
27
| NM_002592.2 | PCNA | Proliferating cell nuclear antigen, transcript variant 1 | -7.25 | 4.32E-04 |
28
| NM_182649.1 | PCNA | Proliferating cell nuclear antigen, transcript variant 2 | -4.53 | 0.0219 |
29
| NM_000321.2 | RB1 | Retinoblastoma 1 | -5.85 | 0.0097 |
DNA repair
| |||||
30
| NM_138292.3 | ATM | Ataxia telangiectasia mutated, transcript variant 2 | 4.49 | 0.0328 |
31
| NM_007306.2 | BRCA1 | Breast cancer 1, early onset, transcript variant BRCA1-exon4 | -3.56 | 0.0371 |
Immunity and defense
| |||||
32
| NM_001024844.1 | CD82 | CD82 molecule, transcript variant 2 | -4.80 | 0.0127 |
33
| NM_002996.3 | CX3CL1 | Chemokine (C-X3-C motif) ligand 1 | 9.44 | 2.67E-05 |
34
| NM_153201.1 | HSPA8 | Heat shock 70kDa protein 8, transcript variant 2 | -3.47 | 0.0350 |
35
| NM_006597.3 | HSPA8 | Heat shock 70kDa protein 8, transcript variant 1 | -3.31 | 0.0100 |
36
| NM_001098631.1 | IRF5 | Interferon regulatory factor 5, transcript variant 7 | 13.39 | 3.78E-04 |
37
| NM_182826.1 | SCARA3 | Scavenger receptor class A, member 3, transcript variant 2 | 6.60 | 0.0139 |
38
| NM_003246.2 | THBS1 | Thrombospondin 1 | 81.54 | 1.31E-08 |
39
| NM_003247.2 | THBS2 | Thrombospondin 2 | 4.16 | 0.0408 |
Metabolism
| |||||
40
| NM_020128.1 | MDM1 | Mdm4, transformed 3T3 cell double minute 1, transcript variant 2 | -3.98 | 0.0146 |
41
| NM_020128.1 | MDM1 | Mdm4, transformed 3T3 cell double minute 1, transcript variant 2 | 7.64 | 0.0023 |
42
| NM_004530.2 | MMP2 | Matrix metallopeptidase 2 | -12.58 | 0.0017 |
43
| NM_000603.3 | NOS3 | Nitric oxide synthase 3 | 11.77 | 4.62E-05 |
44
| NM_033239.2 | PML | Promyelocytic leukemia, transcript variant 9 | -6.24 | 2.81E-04 |
45
| NM_001034.1 | RRM2 | Ribonucleotide reductase M2 polypeptide | -3.38 | 0.0476 |
46
| NM_005063.4 | SCD | Stearoyl-CoA desaturase | -4.04 | 0.0039 |
Transcription regulation
| |||||
47
| NM_006210.1 | PEG3 | Paternally expressed 3 | 42.85 | 2.28E-05 |
48
| NM_003068.3 | SNAI2/SLUG | Snail homolog 2 | -27.69 | 9.45E-07 |
49
| NM_004295.3 | TRAF4 | TNF receptor-associated factor 4 | 3.75 | 0.0033 |
50
| NM_152240.1 | ZMAT3/WIG1 | Zinc finger, matrin type 3, transcript variant 2 | -6.97 | 0.0024 |
Signal transduction
| |||||
51
| NM_014376.2 | CYFIP2 | Cytoplasmic FMR1 interacting protein 2, transcript variant 3 | 17.86 | 1.20E-08 |
52
| NM_004431.2 | EPHA2 | EPH receptor A2 | 5.42 | 0.0031 |
53
| NM_001005914.1 | SEMA3B | Semaphorin 3B, transcript variant 2 | 156.75 | 1.59E-08 |
Unknown function
| |||||
54
| NM_012242.2 | DKK1 | Dickkopf homolog 1 | -17.41 | 0.0097 |
55
| NM_182915.2 | STEAP3 | STEAP family member 3, transcript variant 1 | 8.22 | 0.0024 |
56
| NM_005802.2 | TOPORS | Topoisomerase I binding, arginine/serine-rich | -4.13 | 0.0350 |
Transcript expression of P53 target genes in melanoma cell lines
No. | Accession No. | Gene Symbol | Gene Name | Fold change | p-value |
---|---|---|---|---|---|
Apoptosis
| |||||
Induction of apoptosis
| |||||
1
| NM_138765.2 | BAX | BCL2-associated X protein, transcript variant sigma | -4.19 | 0.0062 |
2
| NM_004324.3 | BAX
1
| BCL2-associated X protein, transcript variant beta | -4.67 | 0.0059 |
3
| NM_001225.3 | CASP4 | Caspase 4, transcript variant alpha | -2.32 | 0.0027 |
4
| NM_001226.3 | CASP6
1
| Caspase 6, transcript variant alpha | 3.17 | 0.0401 |
5
| NM_021202.1 | TP53INP2 | Tumor protein p53 inducible nuclear protein 2 | -3.02 | 0.0034 |
6
| NM_147184.1 | TP53I3/PIG3 | Tumor protein p53 inducible protein 3, transcript variant 2 | -3.67 | 0.0031 |
Inhibition of apoptosis
| |||||
7
| NM_138578.1 | BCL2L1
1
| B-cell CLL/lymphoma 2-like 1, transcript variant 1 | -2.95 | 0.0124 |
8
| NM_003840.3 | TNFRSF10D
1
| Tumor necrosis factor receptor superfamily, member 10d | -33.80 | 0.0005 |
Cell cycle, proliferation and differentiation
| |||||
9
| NM_001012271.1 | BIRC5 | Baculoviral IAP repeat-containing 5 (Survivin), transcript variant 3 | 4.60 | 0.0024 |
BIRC5
|
12.61
|
1.8E-05
| |||
10
| NM_031966.2 | CCNB1 | Cyclin B1 | 18.17 | 0.0005 |
11
| NM_001759.2 | CCND2
1
| Cyclin D2 | -75.73 | 0.0427 |
12
| NM_001238.1 | CCNE1
1
| Cyclin E1, transcript variant 1 | 2.34 | 0.0310 |
13
| NM_057735.1 | CCNE2 | Cyclin E2, transcript variant 2 | 9.33 | 0.0198 |
14
| NM_001786.2 | CDC2/CDK1 | Cell division cycle 2, transcript variant 1 | 6.32 | 0.0127 |
15
| NM_001790.3 | CDC25C | Cell division cycle 25 homolog C, transcript variant 1 | 17.05 | 0.0020 |
16
| NM_001790.3 | CDC25C | Cell division cycle 25 homolog C, transcript variant 1 | 15.26 | 0.0001 |
17
| NM_022809.2 | CDC25C | Cell division cycle 25 homolog C, transcript variant 2 | 13.39 | 0.0011 |
CDC25C
|
18.21
|
0.0004
| |||
18
| NM_058197.3 | CDKN2A
1
| Cyclin-dependent kinase inhibitor 2A, transcript variant 3 | 3.14 | 0.0472 |
CDKN2A
|
1.56
|
0.4706
| |||
19
| NM_001274.3 | CHEK1 | CHK1 checkpoint homolog | 3.14 | 0.0105 |
20
| NM_016426.4 | GTSE1 | G-2 and S-phase expressed 1 | 6.69 | 0.0014 |
21
| NM_182649.1 | PCNA
1
| Proliferating cell nuclear antigen, transcript variant 2 | 4.66 | 0.0397 |
22
| NM_006034.2 | TP53I11 | Tumor protein p53 inducible protein 11 | -54.31 | 2.31E-06 |
DNA repair
| |||||
23
| NM_007304.2 | BRCA1 | Breast cancer 1, transcript variant BRCA1-delta11b | 3.70 | 0.0092 |
24
| NM_007299.2 | BRCA1 | Breast cancer 1, transcript variant BRCA1-delta14-17 | 3.06 | 0.0068 |
BRCA1
|
5.47
|
0.0037
| |||
Immunity and defense
| |||||
25
| NM_000963.1 | PTGS2 | Prostaglandin-endoperoxide synthase 2 | -15.59 | 0.0074 |
Metabolism
| |||||
26
| NM_201397.1 | GPX1 | Glutathione peroxidase 1, transcript variant 2 | -2.16 | 0.0011 |
27
| NM_004530.2 | MMP2 | Metallopeptidase 2 | -6.61 | 0.0296 |
28
| NM_001034.1 | RRM2
1
| Ribonucleotide reductase M2 polypeptide | 10.48 | 0.0068 |
Signal Transduction
| |||||
29
| NM_000623.2 | BDKRB2 | Bradykinin receptor B2 | -16.54 | 0.0040 |
Transport
| |||||
30
| NM_002539.1 | ODC1 | Ornithine decarboxylase 1 | 4.12 | 0.0099 |
31
| NM_018976.3 | SLC38A2 | Solute carrier family 38, member 2 | -5.03 | 0.0074 |
Biological function unclassified
| |||||
32
| NM_018685.2 | ANLN | Anillin | 8.88 | 4.17E-06 |
33
| XM_001133677.1 | LOC729264 | PREDICTED: Similar to TP53TG3 protein, transcript variant 2 | 25.41 | 0.0068 |
34
| NM_016212.2 | TP53TG3 | Tumor protein p53 target gene 3 | 102.59 | 0.0005 |
Accession No. | Gene Symbol | Gene Name | Fold change (WT vs MT) | p-value |
---|---|---|---|---|
NM_001012271.1 | BIRC5 | Baculoviral IAP repeat-containing 5 (Survivin), transcript variant 3 | 2.81 | 0.03297 |
BIRC5
|
2.42
|
0.01001
| ||
NM_001080125.1 | CASP8 | Caspase 8, transcript variant G | -5.88 | 0.02569 |
NM_001024844.1 | CD82 | CD82 molecule, transcript variant 2 | -19.52 | 0.03576 |
NM_001790.3 | CDC25C | Cell division cycle 25 homolog C, transcript variant 1 | -2.57 | 0.00007 |
NM_058195.2 | CDKN2A | Cyclin-dependent kinase inhibitor 2A, transcript variant 4 | -88.58 | 0.0159 |
CDKN2A
|
-34.08
|
2.2E-05
| ||
NM_003068.3 | SNAI2/SLUG | Snail homolog 2 | -8.92 | 0.00005 |
NM_005862.2 | STAG1 | Stromal antigen 1 | 2.33 | 0.04536 |
Inhibition of P53 has limited effect on the mRNA expression of known P53 target genes in melanoma
Accession No. | Gene Symbol | Gene Name | Melan. | IgR3 | Mel-RM |
---|---|---|---|---|---|
Genes regulated similarly by p53 KO in melanocytes and melanoma cells
| |||||
Cell cycle, proliferation or differentiation
| |||||
NM_000389.2 | CDKN1A | Cyclin-dependent kinase inhibitor 1A, transcript variant 1 |
2.68
| - |
7.12
|
NM_004864.1 | GDF15 | Growth differentiation factor 15 |
3.42
| - |
6.11
|
Transcription regulation/signal transduction
| |||||
NM_014376.2 | CYFIP2 | Cytoplasmic FMR1 interacting protein 2, transcript variant 3 |
5.11
| - |
8.88
|
Genes regulated by p53 KO in melanoma cells but not in melanocytes
| |||||
Apoptosis
| |||||
NM_138578.1 | BCL2L1
1
| BCL2-like 1 (Bcl-xL), transcript variant 1 | - |
-2.06
|
2.01
|
NM_033294.2 | CASP1 | Caspase 1, transcript variant delta | - | - |
-4.20
|
NM_016479.3 | SHISA5 | Shisa homolog 5 | - |
-2.30
| - |
NM_019058.2 | DDIT4 | DNA-damage-inducible transcript 4 | - |
-2.56
| - |
Cell cycle, proliferation or differentiation
| |||||
NM_078467.1 | CDKN1A | Cyclin-dependent kinase inhibitor 1A, transcript variant 2 | - | - |
7.56
|
NM_000548.3 | TSC2 | Tuberous sclerosis 2, transcript variant 1 | - | - |
3.95
|
Immunity and defense
| |||||
NM_003897.3 | IER3 | Immediate early response 3 | - | - |
2.59
|
Metabolism
| |||||
NM_000603.3 | NOS3 | Nitric oxide synthase 3 | -2.72 |
10.65
| - |
Transcription regulation/signal transduction
| |||||
NM_001037333.1 | CYFIP2 | Cytoplasmic FMR1 interacting protein 2, transcript variant 1 | - | - |
14.63
|
NM_152546.1 | SRFBP1 | Serum response factor binding protein 1 | - | - |
-2.12
|
NM_004559.3 | YBX1 | Y box binding protein 1 | - |
-2.75
| - |
Transport
| |||||
NM_000593.5 | TAP1 | Transporter 1 | - |
2.07
| - |
Unknown function
| |||||
NM_182915.2 | STEAP3 | STEAP family member 3, transcript variant 1 | - |
4.64
|
-2.94
|
Genes regulated by p53 KO in melanocytes but not in melanoma
| |||||
Apoptosis
| |||||
NM_001040619.1 | ATF3 | Activating transcription factor 3, transcript variant 4 |
-5.30
| - | - |
NM_001008925.1 | RCHY1 | Ring finger and CHY zinc finger domain containing 1, transcript variant 2 |
-12.23
| - | - |
NM_005427.1 | TP73 | Tumor protein p73 |
10.54
| - | 2.84 |
Cell cycle, proliferation or differentiation
| |||||
NM_001012271.1 | BIRC5
1
| Baculoviral IAP repeat-containing 5, transcript variant 3 |
-18.95
| - | - |
NM_004701.2 | CCNB2 | Cyclin B2 |
-3.17
| - | - |
NM_057735.1 | CCNE2
1
| Cyclin E2, transcript variant 2 |
-5.73
| - | - |
NM_001786.2 | CDC2
1
| Cell division cycle 2, transcript variant 1 |
-8.42
| - | - |
NM_022809.2 | CDC25C
1
| Cell division cycle 25 homolog C, transcript variant 2 |
-9.40
| - | - |
NM_001259.5 | CDK6 | Cyclin-dependent kinase 6 |
2.15
| - | - |
NM_058195.2 | CDKN2A
1
| Cyclin-dependent kinase inhibitor 2A, transcript variant 4 |
-12.78
| - | - |
NM_006622.2 | PLK2 | Polo-like kinase 2 |
18.00
| -8.19 | - |
NM_014454.1 | SESN1 | Sestrin 1 |
2.69
| - | - |
Immunity and defense
| |||||
NM_003246.2 | THBS1 | Thrombospondin 1 |
-18.25
| - | - |
NM_003247.2 | THBS2 | Thrombospondin 2 |
2.21
| - | - |
NM_033550.3 | TP53RK | TP53 regulating kinase |
-2.08
| - | - |
Transcription regulation/signal transduction
| |||||
NM_000376.2 | VDR | Vitamin D receptor, transcript variant 1 |
-20.88
| - | - |
Inhibition of P53 in melanocytes results in an altered P53 target gene mRNA expression profile that is similar to that observed in melanoma cells
The ability of P53 to regulate genes involved in the cell cycle is significantly reduced in melanoma cells
Biological Process | % of genes regulated by p53 in Melan. (from 673 unique genes) | Over/under | p-value | % of genes regulated by p53 in IgR3 (from 552 unique genes) | Over/under | p-value | % of genes regulated by p53 in Mel-RM (from 380 unique genes) | Over/under | p-value | |
---|---|---|---|---|---|---|---|---|---|---|
1
|
Mitosis
| 7.28 | + | 1.36E-07 |
4.53
|
+
|
5.32E-02
|
5.00
|
+
|
3.80E-02
|
2
| Cellular process | 40.86 | + | 1.55E-07 | 40.40 | + | 5.36E-06 |
41.32
|
+
|
3.07E-05
|
3
|
Cell cycle
| 14.71 | + | 3.28E-06 |
11.78
|
+
|
2.68E-02
|
11.58
|
+
|
7.22E-02
|
4
|
Cytokinesis
| 3.42 | + | 1.05E-05 |
1.45
|
+
|
3.41E-01
|
1.05
|
-
|
5.23E-01
|
5
|
Nucleobase, nucleoside, nucleotide and nucleic acid metabolic process
| 25.85 | + | 1.50E-05 |
19.57
|
+
|
4.33E-01
|
16.05
|
-
|
6.48E-02
|
6
| Metabolic process | 48.89 | + | 6.80E-05 | 47.83 | + | 1.60E-03 |
43.68
|
+
|
2.10E-01
|
7
| Primary metabolic process | 47.25 | + | 6.93E-05 | 45.83 | + | 2.78E-03 |
41.84
|
+
|
2.38E-01
|
8
| Chromosome segregation | 2.82 | + | 1.09E-04 |
0.91
|
-
|
4.93E-01
| 1.84 | + | 1.03E-01 |
9
| Vesicle-mediated transport | 9.06 | + | 5.16E-04 | 9.42 | + | 5.34E-04 | 8.95 | + | 9.33E-03 |
10
| Anatomical structure morphogenesis | 8.62 | + | 1.03E-03 | 8.70 | + | 2.18E-03 | 8.95 | + | 5.71E-03 |
11
| Cellular component morphogenesis | 8.62 | + | 1.03E-03 | 8.70 | + | 2.18E-03 | 8.95 | + | 5.71E-03 |
12
| Ectoderm development | 10.40 | + | 1.26E-03 | 10.14 | + | 5.92E-03 | 11.05 | + | 3.70E-03 |
13
| Localization | 1.63 | + | 1.27E-03 | 1.81 | + | 9.74E-04 | 0.79 | + | 3.35E-01 |
14
| Nervous system development | 9.36 | + | 1.38E-03 | 8.33 | + | 3.56E-02 | 10.00 | + | 3.79E-03 |
15
| Cellular component organization | 10.40 | + | 1.70E-03 | 11.05 | + | 7.71E-04 | 10.26 | + | 1.87E-02 |
16
| Cell-cell signaling | 9.66 | + | 2.12E-03 |
11.96
| + |
4.33E-06
| 7.63 | + | 2.57E-01 |
17
| Lipid metabolic process | 8.32 | + | 2.56E-03 | 9.06 | + | 7.31E-04 | 7.89 | + | 4.00E-02 |
18
| Defense response to bacterium | 1.19 | + | 3.85E-03 | 1.27 | + | 4.74E-03 | 0.53 | + | 4.06E-01 |
19
| System process | 14.41 | + | 5.19E-03 | 14.86 | + | 4.42E-03 | 13.68 | + | 6.96E-02 |
20
| Meiosis | 2.08 | + | 6.73E-03 | 1.27 | + | 2.86E-01 | 2.11 | + | 3.29E-02 |