Supplemental data: A.1 Primer sequences used for SYBR Green real time RT-PCR Table A.1: Primer sequences used for quantitative SYBR Green real time RT-PCR for validation of genes putatively differentially expressed in MDA-MB-468 cells depending on HER2 expression. Gene Accession Primer sequence (5´3´) product # HDACfor GACTTCTACCAACCCACGTGCA NM_004964 HDACrev TTCAACGCATTCCCCATGC API5for TCCAGGGTAAAACGGGTGAG NM_006595 API5rev CAACGACTTTAATCTTGTTCTCTTCTGT ANXA4for GCTCTGCTGGCTATAGTAAAGTGCA NM_001153 ANXA4rev CGAGAAACCATCACTCTGATGAGG DTX4for GCATTCAGGGACCGGAACAC XM_166213 DTX4rev CACGAGCAGCAGCTTCAGAACTT HIF1Afor GATGAAAGAATTACCGAATTGATGG NM_181054 HIF1Arev GTCCTGTGGTGACTTGTCCTTTAGT NR4A2for CGCGGAGAACTCCTAAGGAGG NM_173171 NR4A2rev AACACAAGGCATGGCTTCAGC ZNF165for CCTATGGGACTGTGATAATGAGAGTG NM_003447 ZNF165rev CCAGCTTTGGCATGGATCTACT P4HBfor TCCTCTTTAAGAAGTTTGATGAAGGC NM_000918 P4HBrev CTGTCTGCTCGGTGAACTCGAT TOB1for TGTGTTTGCAGCCTATGGAGG NM_005749 TOB1rev AACAGGCTGGAATTGCTGGTTA SPINT2for GTGCCACTGTCACAGAGAATGC NM_021102 SPINT2rev TTCTGGGAGCACTTGGGACA BHLHB2for CACGGAGACCTACCAGGGATG NM_003670 BHLHB2rev TTGTAGGTCTCCTTGCTGTCCTC BCL2L2for GATGGTGGCCTACCTGGAGA NM_004050 BCL2L2rev AGAGCTGTGAACTCCGCCCA RARRES1for ACTTCTTCAACTTCCGGTCCG NM_206963 RARRES1rev CACCTTCCTGAAGTAAAGACTCTGG SCDfor GCTTCGAAACCGCAGTCCTC NM_005063 SCDrev TAGGAGCTAGAGATATCGTCCTGCA ELF3for GCCATGAGGTACTACTACAAACGG NM_004433 ELF3rev ACTCTGGAGAACCTCTTCCTCCT CHD3for AGGAAGACCAAGACAACCAGTCAG NM_005852 CHD3rev TGACTGTCTACGCCCTTCAGGA S100A14for kind gift of Dr. C. Kerkhoff, Experimental NM_020672 S100A14rev Dermatology, University of Münster MARK3SVfor GGAATATGCAAGTGGAGGTGAAGTA NM_002376 MARK3SVrev GTCTGTCCAGCTTGACAACCCT A.2 Antibodies Table A.2: Sources, pre-treatment, conditions and the dilutions for TOB, pTOB, and Epidermal Growth Factor (EGF) Antibody Source Clone Pre-treatment Dilution TOB IBL 4B1 Steamer, 1:25 citrate buffer 30´, pH 6.0 pTOB IBL polyclonal Steamer, 1:5 citrate buffer 30´, pH 6.0 EGF Acris polyclonal Steamer, TRS 1:20 buffer 30´, pH 9.0 A.3 Lists of gene products found to be differentially expressed in HER2 overexpressing MDA-MB-468 cells compared to control cells Table A.3: List of 109 genes found to be upregulated in MDA-MB-468 cells overexpressing HER2 compared to the control cells containing the negative control plasmid vector. Of the 144 sequenced clones 23 could not be unambiguously identified and 12 sequences turned out to be doublets of clones identified before. Genes upregulated in MDA-MB-468 -HER2 cDNA Accession# Locus 1 C219-reactive peptide BC047116 1 2 TARDBP: TAR DNA binding protein NM_007375 1p36.22 PHTF1: putative homeodomain transcription 3 factor, a gene for a protein similar to BAI1- AL133517 1p12-13.3 associated protein 4 D1S155E: NRAS-related gene NM_007158 1p22 KHDRBS1: KH domain containing, RNA 5 binding, signal transduction associated 1 (p62, NM_006559 1p32 Sam68) 6 RPS8: ribosomal protein S8 NM_001012 1p34.1-p32 7 PUM1: pumilio homolog 1 (Drosophila) NM_014676 1p35.2 8 ADIPOR1: Adiponectin receptor 1 NM_015999.2 1p36.13-q41 9 ENO1: enolase 1, (alpha) NM_001428 1p36.3-p36.2 S100A8: S100 calcium binding protein A8 10 NM_002964 1q21 (calgranulin A) 11 S100A14: S100 calcium binding protein A14 NM_020672.1 1q21.1 12 FLJ12671 NM_030980 1q22 13 IPO9: importin 9 NM_018085.3 1q31.3 14 CSRP1: cysteine and glycine-rich protein 1 NM_004078.1 1q32 PR1: voltage-dependent calcium channel 15 NM_183240.1 2q14.1 gamma subunit-like protein 16 hypothetical protein LOC130574 NM_194317 2q23.3 IDH1: isocitrate dehydrogenase 1 (NADP+), 17 NM_005896.2 2q33.3 soluble ACTR8: ARP8 actin-related protein 8 homolog 18 NM_022899.3 3 (yeast) 19 FLJ10618 Hypothetical protein NM_018155 3 20 RYBP: RING1 and YY1 binding protein NM_012234.3 3p14.1 IMPDH2: IMP (inosine monophosphate) 21 NM_000884 3p21.2 dehydrogenase 2 UQCRC1: ubiquinol-cytochrome c reductase 22 NM_003365 3p21.3 core protein I 23 HSPC016 NM_015933.2 3p21.31 24 RPN1: ribophorin I NM_002950.2 3q21.3-q25.2 Table A.1, continued TM4SF1: transmembrane 4 superfamily 25 NM_014220.1 3q21-q25 member 1 GALNT7: UDP-N-acetyl-alpha-D- 26 galactosamine:polypeptide N- NM_017423.1 4q31.1 acetylgalactosaminyltransferase 7 (GalNAc-T7) 27 AC020894 5 28 AC094101 5 29 AC008450 5 CCT5: chaperonin containing TCP1, subunit 5 30 NM_012073 5p15.2 (epsilon) 31 KIBRA: KIBRA protein NM_015238.1 5p35.1 32 CAST: calpastatin (TV2) NM_173060.1 5q15-q21 33 HDAC3: histone deacetylase 3 NM_003883.2 5q31 34 HTGN29: HTGN29 protein NM_020199 5q31.1 35 CSNK1A1: casein kinase 1, alpha 1 NM_001892.3 5q32 36 DKFZP566C243 AK021433.1 6p21.1 37 RDBP: RD RNA binding protein NM_002904.4 6p21.3 WBSCR22: Williams Beuren syndrome chr 38 NM_017528 7 region 22 39 DKFZP564K0822 NM_030796.2 7p11.2 EIF4H: eukaryotic translation initiation factor 4H 40 isoform / WBSCR1: Williams-Beuren syndrome NM_022170 7q11.23 chromosome region 1 41 NADH dehydrogenase (ubiquinone) 1 alpha NM_005000 7q32 subcomplex, 5, 13kDa (NDUFA5) 42 AC023102 8 UBE2V2: ubiquitin-conjugating enzyme E2 43 NM_003350.2 8q11.1 variant 2 POLR2K: polymerase (RNA) II (DNA directed) 44 NM_005034.2 8q22.2 polypeptide K, 7.0kDa 45 TIEG: TGFB inducible early growth response NM_005655 8q22.2 LY6E: lymphocyte antigen 6 complex, locus E 46 NM_002346 8q24.3 (retinoic acid induced gene E) 47 RPS6: ribosomal protein S6 NM_001010 9p21 48 FLJ12643 AK022705 9q21.31 TLE1: transducin-like enhancer of split 1 49 NM_005077.3 9q21.32 (E(sp1) homolog, Drosophila) 50 FBXW2: F-box and WD-40 domain protein 2 NM_012164 9q34 MAN1B1: mannosidase, alpha, class 1B, 51 NM_007230 9q34 member 1 SH3GLB2: SH3-domain GRB2-like endophilin 52 NM_020145.2 9q34 B2 53 AL132656 10 GHITM: growth hormone inducible 54 NM_014394 10q23.1 transmembrane protein 55 EHF: Ets homologous factor NM_012153.3 11 56 API5: apoptosis inhibitor 5 NM_006595.2 11p12-q12 Table A.1, continued 57 AP002812 11q PORIMIN: pro-oncosis receptor inducing 58 NM_052932.1 11q22.1 membrane injury gene MGST1: microsomal glutathione S-transferase 59 NM_020300 12p12.3-p12.1 1 60 CD9: CD9 antigen (p24) NM_001769.2 12p13.3 NET-5: transmembrane 4 superfamily member 61 NM_006675.2 12p13.33 tetraspan NET-5 HNRPA1: heterogeneous nuclear 62 NM_002136.1 12q13.1 ribonucleoprotein A1 63 K-ALPHA-1: tubulin, alpha, ubiquitous NM_006082 12q13.11 MYL6: myosin, light polypeptide 6, alkali, 64 NM_021019.2 12q13.13 smooth muscle and non-muscle CCT2: chaperonin containing TCP1, subunit 2 65 NM_006431.1 12q14.3 (beta) 66 TFDP1: transcription factor Dp-1gene AF550129 13 67 AL390754 13 68 FLJ11305 NM_018386.1 13q34 69 ING1: inhibitor of growth family, member 1 NM_005537 13q34 OXA1L: oxidase (cytochrome c) assembly 1- 70 NM_005015 14q11.2 like 71 AC026583 15 72 CCNDBP1: cyclin D-type binding-protein 1 NM_012142 15q14-q15 73 hypothetical protein FLJ10036 NM_017975 15q22.2 PSMA4: proteasome (prosome, macropain) 74 NM_002789.3 15q24.1 subunit, alpha type, 4 KIF22: kinesin family member 22 / MAZ: MYC- 75 associated zinc finger protein (purine-binding NM_007317.1/NM_002383.1 16p11.2 transcription factor) 76 LITAF: lipopolysaccharide-induced TNF factor NM_004862.1 16p13.3-p12 77 FLJ13154 NM_024598.2 16q13 CES1: carboxylesterase 1 78 NM_001266.3 16q13-q22.1 (monocyte/macrophage serine esterase 1) APPBP1: amyloid beta precursor protein 79 NM_003905 16q22 binding protein 1, 59kDa 80 NQO1: NAD(P)H dehydrogenase, quinone 1 NM_000903 16q22.1 81 UBB: ubiquitin B NM_018955.2 17p12-p11.2 EIF4A1: eukaryotic translation initiation factor 82 NM_001416 17p13 4A, isoform 1 SLC25A11: solute carrier family 25 83 (mitochondrial carrier; oxoglutarate carrier), NM_003562.3 17p13.3 member 11 ERBB2: v-erb-b2 erythroblastic leukemia viral 84 oncogene homolog 2, neuro/glioblastoma NM_004448.2 17q21.1 derived oncogene homolog (avian) 85 SP6: Sp6 transcription factor NM_199262.2 17q21.3-q22 NDUFV2: NADH dehydrogenase (ubiquinone) 86 NM_021074.1 18p11.31-p11.2 flavoprotein 2, 24kDa Table A.1, continued 87 CN2: cytosolic nonspecific dipeptidase NM_018235 18q22.3 PAFAH1B3: platelet-activating factor 88 acetylhydrolase, isoform Ib, gamma subunit NM_002573.2 19q13.1 29kDa SPINT2: serine protease inhibitor, Kunitz type, 89 NM_021102.1 19q13.1 2 (placental bikunin) 90 RPS19: ribosomal protein S19 NM_001022.3 19q13.2 ITPA: inosine triphosphatase (nucleoside 91 NM_033453.2 20p triphosphate pyrophosphatase) 92 TRIB3: tribbles homolog 3 (Drosophila) NM_021158.3 20p12.2-p13 93 PCNA: proliferating cell nuclear antigen NM_002592.2 20pter-p12 94 MANBAL: mannosidase, beta A, lysosomal-like NM_022077.3 20q11.23-q12 DPM1: dolichyl-phosphate 95 mannosyltransferase polypeptide 1, catalytic NM_003859.1 20q13.13 subunit 96 cDNA FLJ90800 fis AK075281 21q22.1 GART: phosphoribosylglycinamide formyl- transferase, phosphoribosylglycinamide 97 NM_000402 21q22.11 synthetase, phosphoribosylaminoimidazole synthetase 98 DSCR3: Down syndrome critical region gene 3 NM_006052.1 21q22.2 99 ZNF278: zinc finger protein 278 NM_014323 22q12.2 100 BX293995 X 101 SMS: spermine synthase NM_004595 Xp22.1 102 PRDX4: peroxiredoxin 4 NM_006406.1 Xp22.13 103 TMSB4X: thymosin, beta 4, X chromosome NM_021109.2 Xq21.3-q22 104 MORF4L2: mortality factor 4 like 2 NM_012286.1 Xq22 105 Z83819 Xq22 TCEAL5:Transcription elongation factor A(SII)- 106 NM_032926.2 Xq22.1 like5 PSMD10: proteasome (prosome, macropain) 107 NM_002814.2 Xq22.3 26S subunit, non-ATPase, 10 108 G6PD: glucose-6-phosphate dehydrogenase NM_000402 Xq28 109 cDNA FLJ25150 fis AK057879 Table A.4: List of 112 genes found to be upregulated in MDA-MB-468 cells overexpressing HER2 compared to the control cells