Genome-wide approach to identify transcripts induced by DNA demethylation in tumor cells Axelle Loriot Computational Biology and Bioinformatics de Duve Institute
Methylation profiles are profoundly altered in cancer cells 5m Cytosine Cytosine SOMATIC CELL HYPERMETYLATION HYPOMETYLATION TUMOR CELL TUMOR SUPPRESSOR GENES GERMLINE SPECIFIC GENES
Genome-wide approach to uncover the set of transcripts activated by DNA demethylation in tumors Ø Other tissue specific genes reactivated by demethylation in tumors ? Ø Tumor specific transcripts could be isoforms using demethylated alternative promoters Other tissue specific genes? TUMOR CELL GERMLINE SPECIFIC GENES
METHODOLOGY Bisulfite-seq RNA-seq fastq fastq OMICS CATALOGUE OF LUNG ADENOCARCINOMA CELL LINES hisat2 Accession: PRJDB2256 BisMark 26 LUAD cell lines de novo transcript StringTie assembling Transcripts activated in cell lines showing an inverse correlation between methylation and expression Normal tissues Ø Tissue specificity analysis Ø Methylation profile analysis
214 transcripts identified Reference gene No gene reference Normal 128 transcripts Alternative 63 transcripts Unknown 23 transcripts
AGO1 : an example of alternative transcript activated by demethylation chr1. T lung sperm Normal tissues testis esophagus skin keratinocytes adipose colon liver Normal pancreas stomach 128 intestine Normal tissues transcripts placenta thyroid cortex heart k fibroblasts A427 A549 ABC1 H1437 H1648 H1703 Tumor cell lines H1819 H1975 H2126 H2347 H322 LC2ad PC9 RERF_LC_Ad1 Alternative RERF_LC_Ad2 63 RERF_LC_KJ RERF_LC_OK VMRC_LCD transcripts H2228 II18 PC7 H1650 PC3 RERF_LC_MS H1299 PC14 Tumor cell lines RERF_LC_Ad1 Unknown RERF_LC_Ad2 23 RERF_LC_KJ RERF_LC_OK transcripts RERF_LC_MS
Tissue specificity Reference gene Normal RNAseq of 53 normal tissues 128 transcripts Multiple samples per tissues Processed data only => Expression values are given at Gene level Alternative 63 transcripts Unknown 23 transcripts
Tissue specificity LUAD cell lines Normal tissues (GTEx) AS1 log1p(TPM) SLC22A18AS GDPD4 6 TDRD1 DNAH8 CCDC33 C9ORF84 LINC01511 4 MAGEA1 DDX53 MAGEA11 FTHL17 NAA11 2 MAGEC1 LINC01518 PASD1 MAGEA4 AC074389.2 0 RNF17 INSL6 MAGEC2 MAGEB2 matrix_4 DSCR8 TAF7L PRAME FMR1NB BRDT AC109635.4 COX7B2 CT83 AC063977.6 CCDC144NL ST8SIA6 − AS1 AC010168.1 LINC00689 CCDC144NL − AS1 CSAG1 AC090502.1 AP002383.3 AL160408.1 POTEF AC092506.1 FAM9C ZBTB46 − AS1 ANKRD62 Z98257.1 LINC00200 AC106771.1 AC100839.1 AC023824.3 AC087269.1 AC109635.2 LINC02492 AC097374.1 BAGE2 MKRN9P AC099520.1 MAGEA12 LIN28B BCAR4 INSL4 AC010894.3 PLCE1 − AS1 MUC13 EPS8L3 PHGR1 ARL14 BCL2L15 DQX1 TFF1 HNF4A VIL1 SLC39A5 HGD DRAIC HAVCR1 log1p(TPM) HNF1A − AS1 HNF1A SLC22A18AS ABCC2 NPC1L1 ANGPTL3 XDH FAM25A ADGRF4 ABCA12 ZNF812P FAM83A EPGN TPRXL GPR78 KRT16 LY6D SERPINB5 ALDH3B2 DSG3 GPR87 GJB5 A2ML1 TNS4 MIR205HG C5ORF17 IGBP1P4 matrix_4 LINC01115 AC022733.1 LINC00355 LINC01194 LINC01446 DSCR4 AL929601.1 AC090809.1 LINC02008 LINC02438 MIR548XHG RPS4XP8 AC012363.1 BBOX1 − AS1 FILNC1 CASC19 AL117329.1 CLDN10 − AS1 TRPM2 − AS1 LINC02526 AL109741.2 ZNF556 MKRN3 LINC00460 PNLDC1 TRIM55 KCTD4 Gr MSTRG.26453.1 H322 H1648 PC9 RERF_LC_Ad2 H1650 RERF_LC_KJ H1437 H2126 LC2ad H2347 RERF_LC_Ad1 H1975 II18 H2228 ABC1 A549 PC3 H1703 H1819 H1299 A427 RERF_LC_MS RERF_LC_OK PC14 VMRC_LCD PC7 Lung Testis Adipose_subcut Adipose_visc Spleen Ovary Fallopian_tube Uterus Whole_blood Muscle Heart_2 Heart_1 Artery_tibial Artery_aorta Artery_coronary Brain_1 Brain_2 Brain_3 Brain_6 Brain_7 Brain_8 Brain_9 Brain_10 Brain_11 Brain_12 Brain_13 Brain_4 Brain_5 Pituitary Adrenal_gland Nerve Thyroid Colon_sigmoid Colon_transv Small_intestine Stomach Kidney_cortex Kidney_medulla Liver Pancreas Bladder Prostate Salivary_gland Breast Skin_1 Skin_2 Vagina Esophagus_mucosa Esophagus_gastr_junct Esophagus_musc Group 26 LUAD cell lines 50 normal tissues from Gtex
Tissue specificity LUAD cell lines Normal tissues (GTEx) AS1 log1p(TPM) SLC22A18AS GDPD4 6 TDRD1 DNAH8 CCDC33 C9ORF84 LINC01511 4 MAGEA1 DDX53 MAGEA11 FTHL17 NAA11 2 MAGEC1 LINC01518 PASD1 MAGEA4 AC074389.2 0 RNF17 INSL6 MAGEC2 MAGEB2 matrix_4 DSCR8 TAF7L PRAME FMR1NB BRDT AC109635.4 COX7B2 CT83 AC063977.6 CCDC144NL ST8SIA6 − AS1 AC010168.1 LINC00689 CCDC144NL − AS1 CSAG1 AC090502.1 AP002383.3 AL160408.1 POTEF AC092506.1 FAM9C ZBTB46 − AS1 ANKRD62 Z98257.1 LINC00200 AC106771.1 AC100839.1 AC023824.3 AC087269.1 AC109635.2 LINC02492 AC097374.1 BAGE2 MKRN9P AC099520.1 MAGEA12 LIN28B BCAR4 INSL4 AC010894.3 PLCE1 − AS1 MUC13 EPS8L3 PHGR1 ARL14 BCL2L15 DQX1 TFF1 HNF4A VIL1 SLC39A5 HGD DRAIC HAVCR1 log1p(TPM) HNF1A − AS1 HNF1A SLC22A18AS ABCC2 NPC1L1 ANGPTL3 XDH FAM25A ADGRF4 ABCA12 ZNF812P FAM83A EPGN TPRXL GPR78 KRT16 LY6D SERPINB5 ALDH3B2 DSG3 GPR87 GJB5 A2ML1 TNS4 MIR205HG C5ORF17 IGBP1P4 matrix_4 LINC01115 AC022733.1 LINC00355 LINC01194 LINC01446 DSCR4 AL929601.1 AC090809.1 LINC02008 LINC02438 MIR548XHG RPS4XP8 AC012363.1 BBOX1 − AS1 FILNC1 CASC19 AL117329.1 CLDN10 − AS1 TRPM2 − AS1 LINC02526 AL109741.2 ZNF556 MKRN3 LINC00460 PNLDC1 TRIM55 KCTD4 Gr MSTRG.26453.1 H322 H1648 PC9 RERF_LC_Ad2 H1650 RERF_LC_KJ H1437 H2126 LC2ad H2347 RERF_LC_Ad1 H1975 II18 H2228 ABC1 A549 PC3 H1703 H1819 H1299 A427 RERF_LC_MS RERF_LC_OK PC14 VMRC_LCD PC7 Lung Testis Adipose_subcut Adipose_visc Spleen Ovary Fallopian_tube Uterus Whole_blood Muscle Heart_2 Heart_1 Artery_tibial Artery_aorta Artery_coronary Brain_1 Brain_2 Brain_3 Brain_6 Brain_7 Brain_8 Brain_9 Brain_10 Brain_11 Brain_12 Brain_13 Brain_4 Brain_5 Pituitary Adrenal_gland Nerve Thyroid Colon_sigmoid Colon_transv Small_intestine Stomach Kidney_cortex Kidney_medulla Liver Pancreas Bladder Prostate Salivary_gland Breast Skin_1 Skin_2 Vagina Esophagus_mucosa Esophagus_gastr_junct Esophagus_musc Group 26 LUAD cell lines 50 normal tissues from Gtex Lung
Tissue specificity LUAD cell lines Normal tissues (GTEx) AS1 log1p(TPM) SLC22A18AS GDPD4 6 TDRD1 DNAH8 CCDC33 C9ORF84 LINC01511 4 MAGEA1 DDX53 MAGEA11 FTHL17 NAA11 2 MAGEC1 LINC01518 PASD1 MAGEA4 AC074389.2 0 RNF17 INSL6 MAGEC2 MAGEB2 matrix_4 DSCR8 Testis specific genes TAF7L PRAME FMR1NB BRDT AC109635.4 COX7B2 CT83 AC063977.6 CCDC144NL ST8SIA6 − AS1 AC010168.1 LINC00689 CCDC144NL − AS1 CSAG1 AC090502.1 AP002383.3 AL160408.1 POTEF AC092506.1 FAM9C ZBTB46 − AS1 ANKRD62 Z98257.1 LINC00200 AC106771.1 AC100839.1 AC023824.3 AC087269.1 AC109635.2 LINC02492 AC097374.1 BAGE2 MKRN9P AC099520.1 MAGEA12 LIN28B BCAR4 INSL4 AC010894.3 PLCE1 − AS1 MUC13 EPS8L3 PHGR1 ARL14 BCL2L15 DQX1 TFF1 HNF4A VIL1 SLC39A5 HGD DRAIC HAVCR1 log1p(TPM) HNF1A − AS1 HNF1A SLC22A18AS ABCC2 NPC1L1 ANGPTL3 XDH FAM25A ADGRF4 ABCA12 ZNF812P FAM83A EPGN TPRXL GPR78 KRT16 LY6D SERPINB5 ALDH3B2 DSG3 GPR87 GJB5 A2ML1 TNS4 MIR205HG C5ORF17 IGBP1P4 matrix_4 LINC01115 AC022733.1 LINC00355 LINC01194 LINC01446 DSCR4 AL929601.1 AC090809.1 LINC02008 LINC02438 MIR548XHG RPS4XP8 AC012363.1 BBOX1 − AS1 FILNC1 CASC19 AL117329.1 CLDN10 − AS1 TRPM2 − AS1 LINC02526 AL109741.2 ZNF556 MKRN3 LINC00460 PNLDC1 TRIM55 KCTD4 Gr MSTRG.26453.1 H322 H1648 PC9 RERF_LC_Ad2 H1650 RERF_LC_KJ H1437 H2126 LC2ad H2347 RERF_LC_Ad1 H1975 II18 H2228 ABC1 A549 PC3 H1703 H1819 H1299 A427 RERF_LC_MS RERF_LC_OK PC14 VMRC_LCD PC7 Lung Testis Adipose_subcut Adipose_visc Spleen Ovary Fallopian_tube Uterus Whole_blood Muscle Heart_2 Heart_1 Artery_tibial Artery_aorta Artery_coronary Brain_1 Brain_2 Brain_3 Brain_6 Brain_7 Brain_8 Brain_9 Brain_10 Brain_11 Brain_12 Brain_13 Brain_4 Brain_5 Pituitary Adrenal_gland Nerve Thyroid Colon_sigmoid Colon_transv Small_intestine Stomach Kidney_cortex Kidney_medulla Liver Pancreas Bladder Prostate Salivary_gland Breast Skin_1 Skin_2 Vagina Esophagus_mucosa Esophagus_gastr_junct Esophagus_musc Group 26 LUAD cell lines 50 normal tissues from Gtex Lung Testis
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