As previously described[2], cultured cells were cross-linked with 1% ... Subramanian, A., et al., GSEA-P: a desktop application for Gene Set Enrichment. Analysis.
Cell Reports Supplemental Information
LncRNA HOTAIR Enhances the AndrogenReceptor-Mediated Transcriptional Program and Drives Castration-Resistant Prostate Cancer Ali Zhang, Jonathan C. Zhao, Jung Kim, Ka-wing Fong, Yeqing Angela Yang, Debabrata Chakravarti, Yin-Yuan Mo, and Jindan Yu
Supplemental Experimental Procedures Plasmids and Reagents
Full length (1-2337), 5’ (1-360), 5’ (1-1550), 3’ (1550-2337), 3’ (660-2337) and anti-sense full length HOTAIR fragment were cloned into NotI site of pcDNA3 vector and were used as template to synthesize RNA probes by T7 polymerase. Plasmids expressing AR deletion mutants and MDM2 were generated by PCR and are all FLAG-tagged. HA-Ub plasmid comes from Addgene. All PCR primers for cloning were listed in Supplementary information table S4 and high fidelity enzyme Phusion was used for PCR amplification. All PCR products were verified by DNA sequencing. Specific antibodies used in our work include: rabbit AR (PG21, Millipore), rabbit AR (SC815x, Santa Cruz), mouse AR (sc-7305, Santa Cruz), mouse EZH2 (612667, B.D.), rabbit FLAG (F7425, Sigma), mouse FLAG (F1804-50UG, sigma), rabbit HA (ab9110, Abcam), mouse GAPDH (ab9448, Abcam), mouse tubulin (sc-32293, Santa Cruz), and rabbit H3 (ab9448, Abcam). Other chemicals utilized in clude Cycloheximide (C1988-1G, Sigma), MG132 (C2251, Sigma), MDV3100 (S1250, Sellech Chem). For RNA decay assay, 5ug/ml Actinomycin-D (Life Technologies) was used to treat LNCaP cells, and then RNA was collected at 0, 1, 2, 4, 6 hours for subsequent RT-PCR to determine HOTAIR level.
RNA pulldown assay RNA pulldown protocol was described previously [1]. Biotin-labeled RNAs were transcribed with Biotin-UTP,NTP mixture and T7 RNA polymerase (Promega), treated with RNase-free DNase I (promega) and purified with RNeasy Mini kit (QIAGEN). 107 cell pellets were resuspended in 2 ml PBS, 2 ml nuclear isolation buffer (1.28 M sucrose; 40 mM Tris-HCl pH 7.5; 20 mM MgCl2; 4% Triton X-100), and 6 ml water on ice for 20 min. Nuclei were pelleted by centrifugation at 2,500 G for 15min. Nuclear pellet was re-suspended in 1 ml RIP buffer (150 mM KCl, 25 mM Tris pH 7.4, 0.5mM DTT, 0.5% NP40, 1 mM PMSF and protease inhibitor (Roche Complete Protease Inhibitor Cocktail Tablets). Re-suspended nuclei were mechanically sheared using a dounce homogenizer with 15–20 strokes then centrifuge at 13,000 RPM for 10 min. For precipitation assays, the reaction (RNA probe and protein extract) was incubated at RT for 60 min, and sixty ul washed Streptavidin agarose beads (Invitrogen) were added to each binding reaction and further incubated at RT for one hour. After washing with PBS for 5 times, samples were boiled in SDS buffer, and the retrieved protein was detected by standard western
blot technique. For detecting direct binding of AR with HOTAIR, AR-NT recombinant protein was prepared by sub-cloning PCR8-AR-NT-HAX2 into gateway compatible maltose-binding protein (MBP) destination vector, transformed into Escherichia coli BL21-CodonPlus (DE3)RIPL competent cells (Agilent technology) and induced with 1mM isopropyl β-dthiogalactopyranoside at 37 °C for 3 h. Biotin-labeled RNA probe was incubated with recombinant protein before proceeding to IP process.
RNA immunoprecipitation RIP protocol was described previously with the following modification [1]. Cells at a concentration of 2 million cells/mL were treated with 0.3% formaldehyde in medium for 10min at 37ºC. 1.25 M glycine dissolved in PBS was added to a concentration of 0.125 M, and the sample was incubated for 5 min at room temperature. Cells were then washed twice in cold PBS and pelleted. The pellet was re-suspended in 1 ml of RIPA buffer (50 mM Tris, pH 7.4, 150 mM NaCl, 1 mM EDTA, 0.1% SDS, 1% NP-40, and 0.5% sodium deoxycholate, 0.5mM DTT and 1mM PMSF/cocktail), incubated on ice with frequent vortex for 30 minutes, and the lysate was obtained by centrifugation at 13,000 RPM for 10 min. Incubated 20 ul protein beads with 1ug antibody and rotated for 4 h at 4ºC to get bead and antibody complex. The complex was added to cell lysates and incubated for overnight at 4ºC. After adding RNase-free DNase I (Promega) 37ºC for 15 min and proteinase K at 45 ºC for 45 min. RNA samples were extracted with 1ml TRIZOL. Proteins isolated before proteinase K treatment from the beads were analyzed by western blot analysis. The data of retrieved RNAs is calculated from the subtraction of RT/input ratio and non-RT/input ratio (for each experiment, n=3).
Western blot analysis Cells were lysed in NP40 Cell Lysis Buffer (Invitrogen). Protein concentration was measured using the Bio-Rad protein assay kit. Twenty ug of total protein were taken up in SDS sample buffer, boiled for 10 minutes at 95°C, separated on a 10% SDS-polyacrylamide gel, and transferred to an Amersham Hybond PVDF membrane. Secondary antibodies include a 1:5,000 dilution of horseradish peroxidase (HRP) –conjugated goat-anti-mouse IgG or goat-anti-rabbit IgG secondary antibodies (Santa Cruz). ECL (GE Health care) Western Blotting Detection Systems was used for Western blot detection.
Co-IP Cells with co-expression of AR (tagged with Flag) and HA-Ub were lysed in IP lysis buffer (50mM Tris-Cl pH7.4, 150mM NaCl, 1mM EDTA, 1% Triton X-100, Roche protease inhibitor cocktail). An aliquot of the cell lysate was kept as input for Western blot analysis. Cell lysate was first pre-cleared with protein A/G-agarose beads at 4°C for 2 h. Then the pre-cleared lysate was incubated with rabbit anti-HA (1:2000 ab9110, Ab cam) or mouse IgG (as a negative control) overnight on a rocker platform. The next day, protein A/G-agarose beads were added to the mixture and incubated at 4°C for 2 h. Then beads were pelleted and washed four times with IP lysis buffer before boiling in SDS sample buffer. Western blotting analysis were performed using mouse anti-AR (1:2000, sc-7305, Santa Cruz) according to protocol as described previously.
ChIP As previously described[2], cultured cells were cross-linked with 1% formaldehyde for 10 min and the crosslinking was inactivated by 0.125M glycine for 5 min at room temperature (RT). Cells were then rinsed with cold 1X PBS twice. The following steps were performed at 4°C. Cell pellets were re-suspended and incubated in cell lysis buffer + 10ul/ml PMSF and protease inhibitor (Roche PI) for 10 min. Nuclei pellets were centrifuged down at 5,000 rpm for 5 min, resuspended in nuclear lysis buffer, and then incubated for another 10min. Chromatin was sonicated to an average length of 500bp and then centrifuged at 14,000rmp for 10min to remove the debris. Supernatants containing chromatin fragments were incubated with agarose/protein A or G beads (Upstate) for 15 min and centrifuged at 5,000 rpm for 5min to reduce nonspecific binding. To immunoprecipitated protein/chromatin complexes, the supernatants were incubated with 3-5ug of antibody overnight, then 50ul of agarose/protein A or G beads was added and incubated for 1.5 hour. Beads were washed twice with 1X dialysis buffer and four times with IP wash buffer. The antibody/protein/DNA complexes were eluted with 150ul IP elution buffer twice. To reverse the crosslinks, the complexes were incubated in elution buffer + 10ug RNase A and 0.3 M NaCl at 67°C for 4 hours. DNA/proteins were precipitated with ethanol, air-dried, and dissolved in 100ul of TE. Proteins were then digested by proteinase K at 45°C for 1 hour and DNA was purified with QIAGEN PCR column and eluted with 30ul EB.
ChIP-seq Peak analysis For ChIP-seq, HOMER peak finding algorithm searches for peaks of fixed size to achieve maximum sensitivity. Homer automatically optimized parameters needed for peak finding: the size of the peak and the length of fragments. We set the minimum distance between peaks using 2.5x peak size. The cutoff for statistically significant peaks was selected at a false discovery rate of 0.001 determined by peak finding using randomized tag positions in a genome with an effective size of 2 billion base pair. We required the tag density at peaks to be 4-fold greater than in the surrounding 10 kb region to exclude putative peaks identified in regions of genomic duplication. Only one tag from each unique position was considered to filter clonal signal from the sequencing.
Gene Set Enrichment Analysis (GSEA) Expression profiling data of LNCaP control and HOTAIR in the absence and presence of androgen were generated by Illumina HumanHT-12 v 4.0 Expression BeadChip. GSEA[3] was performed. Gene expression fold changes in experimental cells relative to control were calculated and genes having at least 2 fold changes in HOTAIR treated cells were defined as HOTAIR-regulated gene sets. Differentially expressed genes (at least 2-fold changes) in LNCaP Ctrl cells androgen-treated compared to hormone-deprived were defined as androgen-regulated gene sets.
In situ hybridization ISH was used to detect HOTAIR in clinical specimens [4]. Biotin-labeled antisense HOTAIR LNA probe /5Biosg/G+C+C+TTGCTCCCTT+G+CCTGCATTTCT+C+T+G was synthesized by EXIQON. For paraffin embedded tissue, after de-paraffinization and rehydration, the samples were treated with peroxidase-quenching solution; proteinase K was added to digest tissues before pre-hybridization and hybridization, which were carried out at 56 ºC for30 min and 4 h, respectively. Then streptavidin–HRP was used to react with the bound biotin-labeled probe. The signal was further amplified using TSA amplification kit (Perkin Elmer). Finally, the signal was revealed with Ultra Vision One polymer and AEC chromogen (Thermo Fisher Scientific). The stains were then scored on a three-tiered scoring system, using the following criteria for the three tiered system: 0 = negative; 1 = equivocal/uninterpretable; 2 = weak positive; 3 = strong positive.
Electrophoretic mobility shift assays Electrophoretic mobility shift assays were performed using the LightShift Chemiluminescent EMSA kit from Pierce Biotechnology according to the manufacturer’s standard protocol. Briefly, nuclear extract of AR-DBD or AR-NTD transfected 293 T cells was mixed with Biotin labeled HOTAIR lncRNA through in vitro transcription or Biotin labeled ARE oligo to perform the protein and DNA binding reaction. The reaction product was then run on a pre-cast DNA retardation gel, transferred to nylon membrane, crosslinked and detected by chemiluminescence.
Supplemental References 1. 2. 3. 4.
Tsai, M.C., et al., Long noncoding RNA as modular scaffold of histone modification complexes. Science, 2010. 329(5992): p. 689-93. Yu, J., et al., An integrated network of androgen receptor, polycomb, and TMPRSS2ERG gene fusions in prostate cancer progression. Cancer Cell, 2010. 17(5): p. 443-54. Subramanian, A., et al., GSEA-P: a desktop application for Gene Set Enrichment Analysis. Bioinformatics, 2007. 23(23): p. 3251-3. Zhang, A., et al., The human long non-coding RNA-RoR is a p53 repressor in response to DNA damage. Cell Res, 2013. 23(3): p. 340-50.
A
B
H3K27ac Ethl
chr12:54,355,800-54,362,700 54,356K
H3K27ac DHT
54,358K
54,360K
54,362K
H5 54,362,500
54,361,000
AR R1881
ARE pF2/R2
pF3/R3
0.5 0.4
6
0.3
4
0.2 2
0.1
0
0 PSA
KLK2
1 0.8 0.6 0.4 0.2
OPRK1
MET
HOTAIR
20 18 16 14 12 10 8 6 4 2 0
0
0h
1h
2h
4h
6h
V1
8
EtOH R1881
22 R
R1881
0.6
LNCaP
C4 -2B
EtOH
0.7
10
I
1.2
0 6 12 24 48 1nM R1881 stimulation (hr)
aP -ab l
H
0.8
0 0 6 12 24 48 1nM R1881 stimulation (hr)
aP
0 0 6 12 24 48 1nM R1881 stimulation (hr)
LN C
Enrichment over input
1
LNCaP RNA polII ChIP 12
0.5
2
aP
G
FKBP5 HOTAIR HOTAIR HOTAIR HOTAIR pF/R1 pF/R2 pF/R3 H5
3
C4
0
VCaP
VC
1
4
HOTAIR / GAPDH
2
1.0
VCaP
LA P
3
F
5
LN C
4
LNCaP
16 14 12 10 8 6 4 2 0
HOTAIR/GAPDH
R1881
PSA / GAPDH
EtOH
PSA
E
D
LNCaP AR ChIP
5
HOTAIR//GAPDH
Enrichment over input
6
PSA / GAPDH
C
pF1/R1
Supplemental Figure 1. HOTAIR is directly repressed by androgen receptor (AR) and is up-regulated in CRPC (related to Figure 1). (A). H3K27Ac and AR ChIP-seq around the HOTAIR region. H3K27Ac ChIP-seq (top 2 panels) in ethanol and androgen (DHT)-treated LNCaP cells was obtained through re-analysis of previously published data (GSE51621). AR ChIP-seq showed 3 strong AR binding events at enhancers around the HOTAIR gene (Bottom panel). (B). View of AR binding site around the HOTAIR promoter and the primer pairs for AR ChIPqPCR validation as utilized in Figure 1B. (C). AR ChIP-qPCR in LNCaP cells in the presence and absence of androgen showing AR occupancy at known target genes PSA, FKBP5 (positive controls), HOTAIR promoter regions, and a distal HOTAIR region (H5, negative control). (D). QRT-PCR analysis of PSA expression in LNCaP cells stimulated by 1nM androgen R1881 over a time course. Data was normalized to GAPDH. Data shown was mean ± SEM from triplicate experiments. (E-F). QRT-PCR analysis of PSA and HOTAIR expression in VCaP cells stimulated by 1nM androgen R1881 over a time course. Data was normalized to GAPDH and shown as mean ± SEM from triplicate experiments. (G). RNA pol II ChIP-qPCR showing RNA pol II occupancy at several AR-induced (left) and – repressed genes (right) in the presence and absence of androgen. (H). HOTAIR RNA decay assay in the presence and absence of androgen. LNCaP cells were treated with Actinomycin-D 5ug/ml and then RNA was collected in the indicated time. (I). QRT-PCR analysis of HOTAIR expression in a panel of prostate cancer cell lines. Data was normalized to GAPDH. Data shown was mean ± SEM from triplicate experiments.
Supplemental Figure 2. HOTAIR transcript directly interacts with the AR protein (related to Figure 2). (A) RNA pulldown of HOTAIR-binding proteins in LNCaP cells. Silver staining shows the size of enriched proteins. (B-D) AR RIP was performed in LNCaP cells in the presence of androgen (B) and absence of androgen(C,D) using AR antibody targeting AR N terminal (MILLIPORE, PG21) or C terminal (SC-815x), and analyzed by qRT-PCR to detect HOTAIR RNA enrichment in immunoprecipitated complex. U1 is negative control. (E) EZH2 RIP assay was performed in LNCaP cells and HOTAIR enrichment was detected using qRT-PCR. U1 is negative control. Error bars: mean ± SEM. (F) RNA pulldown assay was carried out in LNCaP cells using different HOTAIR probes and analyzed by qRT-PCR to detect HOTAIR enrichment and confirm RNA pulldown efficiency. (G) Diagram depicting AR deletion constructs used in Figure 2F. (H) EMSA assay showing that HOTAIR RNA does not compete with ARE oligo for binding with AR DBD region.
Supplemental Figure 3. HOTAIR stabilizes the AR protein by blocking MDM2-mediated ubiquitination and AR degradation (related to Figure 3). (A) QRT-PCR analysis of HOTAIR expression in LNCaP_ctrl or LNCaP_HOTAIR cells. Data was normalized to GAPDH. Error bars: mean± SEM. (B) QRT-PCR analysis of AR expression in LNCaP_ctrl or LNCaP_HOTAIR cells. Data was normalized to GAPDH. Error bars: mean± SEM. (C) Cytoplasmic and nuclear AR protein levels in control and HOTAIR-overexpressing LAPC4 cells. Tubulin and H3 are used as cytoplasmic and nuclear protein loading controls, respectively. (D) LAPC4_Ctrl, LAPC4_HOTAIR cells were treated with 10uM CHX for 6 hours and then collected at indicated times for western blotting. (E) C4-2B_shctrl, C4-2B_shHOTAIR, and C4-2B_shHOTAIR+ HOTAIR WT cells were treated with 10uM CHX for 6 hours and then collected at indicated times for western blotting. (F) Co-IP showing AR and MDM2 protein interaction in LNCaP with control or HOTAIR overexpression and C4-2B with control or HOTAIR knockdown.
Supplemental Figure 4. HOTAIR enhances AR transcriptional program (related to Figure 4). (A-B) Genome browser view showing AR enrichment on KLK2 and KLK3 genes in LNCaP_ctrl cells compared with LNCaP_HOTAIR cells in the presence of androgen. (C) QRT-PCR analysis of AR downstream genes in LAPC4 cells with control or HOTAIR overexpression. Gene expression was normalized to GAPDH and shown as mean ± SEM from triplicate experiments.
Supplemental Figure 5. HOTAIR overexpression drives androgen-independent AR activation (related to Figure 5). (A-B) Genome browser view showing AR enrichment on KLK2 and KLK3 in LNCaP_ctrl cells compared with LNCaP_HOTAIR cells in the absence of androgen. (C) QRT-PCR analysis of HOTAIR expression in C4-2B_shctrl or C4-2B_shHOTAIR cells. Data was normalized to GAPDH. Error bars: mean± SEM.
Supplemental Tables Supplemental Table 1 (related to Figure 1). LncRNAs that are bound by AR in LNCaP cells. Supplemental Table 2 (related to Figure 6). GO terms that are enriched by HOTAIR-induced genes in LNCaP cells. Supplemental Table 3 (related to Figure 6). GO terms that are enriched by HOTAIR-repressed genes in LNCaP cells. Supplemental Table 4 (related to Figure 1). Oligonucleotide sequences used in this study.
Table S2: GO terms that are enriched by HOTAIR‐induced genes in LNCaP, Related to Figure 6. Term Category Count % PValue Genes STEAP3, KIF23, MAD1L1, KIFC1, NBN, PRC1, E2F6, PKMYT1, AURKA, PTTG1, AURKB, CCNE2, KIF2C, MLL5, RNF103, OIP5, INCENP, ASPM, CDCA3, STAG1, ANAPC1, CCNH, SGOL1, NUSAP1, RB1, TACC1, NCAPD3, MCM6, RIF1, HAUS8, NUP43, NEK2, TIPIN, BCCIP, ANLN, CHEK2, CEP55, KIAA1009, HJURP, NCAPG2, KATNA1, NPAT, FBXO5, ZWILCH, TERF2, BUB3, DLGAP5, SP_PIR_KEYWORDS cell cycle 60 7.978723 3.54E‐17 NUF2, CENPF, BIRC5, CENPE, CDC20, RACGAP1, CDKN3, CDC25C, SIRT2, CDC25B, RASSF1, KIF20B, CENPV KIF23, MAD1L1, KIFC1, NEK2, TIPIN, ANLN, PTTG1, CEP55, AURKB, KIAA1009, KIF2C, NCAPG2, OIP5, INCENP, KATNA1, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, SGOL1, NUF2, CENPF, NUSAP1, CDC20, BIRC5, CENPE, CDC25C, SP_PIR_KEYWORDS mitosis 37 4.920213 8.88E‐17 SIRT2, NCAPD3, CDC25B, CENPV, KIF20B, HAUS8, NUP43 KIF23, MAD1L1, KIFC1, NEK2, TIPIN, PKMYT1, ANLN, AURKA, PTTG1, CEP55, AURKB, KIAA1009, KIF2C, NCAPG2, OIP5, INCENP, KATNA1, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, DLGAP5, SGOL1, NUF2, CENPF, NUSAP1, CDC20, GO:0007067~mitosis GOTERM_BP_FAT 41 5.452128 1.09E‐16 CENPE, BIRC5, PBK, CDC25C, NCAPD3, SIRT2, CDC25B, CENPV, KIF20B, HAUS8, NUP43 KIF23, MAD1L1, KIFC1, NEK2, TIPIN, PKMYT1, ANLN, AURKA, PTTG1, CEP55, AURKB, KIAA1009, KIF2C, NCAPG2, OIP5, GO:0000280~nuclear INCENP, KATNA1, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, DLGAP5, SGOL1, NUF2, CENPF, NUSAP1, CDC20, division GOTERM_BP_FAT 41 5.452128 1.09E‐16 CENPE, BIRC5, PBK, CDC25C, NCAPD3, SIRT2, CDC25B, CENPV, KIF20B, HAUS8, NUP43 KIF23, ADCY3, KIFC1, MAD1L1, NBN, PRC1, PKMYT1, OSGIN2, AURKA, PTTG1, AURKB, KIF2C, OIP5, INCENP, ASPM, CDCA3, STAG1, ANAPC1, SGOL1, MND1, NUSAP1, RB1, PBK, TACC3, NCAPD3, HAUS8, NUP43, NEK2, TIPIN, ANLN, CEP55, KIAA1009, NCAPG2, KATNA1, MNS1, FBXO5, ZWILCH, BUB3, DLGAP5, NUF2, CENPF, BIRC5, CENPE, CDC20, CDC25C, GO:0000279~M phase GOTERM_BP_FAT 50 6.648936 1.70E‐16 RAD54L, SIRT2, CDC25B, CENPV, KIF20B KIF23, MAD1L1, KIFC1, NEK2, TIPIN, PKMYT1, ANLN, AURKA, PTTG1, CEP55, AURKB, KIAA1009, KIF2C, NCAPG2, OIP5, GO:0000087~M phase INCENP, KATNA1, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, DLGAP5, SGOL1, NUF2, CENPF, NUSAP1, CDC20, of mitotic cell cycle GOTERM_BP_FAT 41 5.452128 2.09E‐16 CENPE, BIRC5, PBK, CDC25C, NCAPD3, SIRT2, CDC25B, CENPV, KIF20B, HAUS8, NUP43 KIF23, MAD1L1, KIFC1, NEK2, TIPIN, PKMYT1, ANLN, AURKA, PTTG1, CEP55, AURKB, KIAA1009, KIF2C, NCAPG2, OIP5, GO:0048285~organelle INCENP, KATNA1, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, DLGAP5, SGOL1, NUF2, CENPF, NUSAP1, CDC20, fission GOTERM_BP_FAT 41 5.452128 5.69E‐16 CENPE, BIRC5, PBK, CDC25C, NCAPD3, SIRT2, CDC25B, CENPV, KIF20B, HAUS8, NUP43
GOTERM_BP_FAT
GO:0022403~cell cycle phase
54 7.180851
GOTERM_BP_FAT
GO:0051301~cell division
44 5.851064
GOTERM_BP_FAT
GO:0000278~mitotic cell cycle
48 6.382979
HGSNAT, KIFC1, LDHA, PDLIM7, PDLIM5, SLC9A2, TUBB2C, INTS1, AURKA, AURKB, ZNF638, PTTG1, KIAA0319L, MAP3K5, MAK16, DHX38, GAB2, AGPS, PLOD2, RAVER2, INTS4, INCENP, PIK3C3, PLS1, VPS13A, FAM103A1, NUP35, KIAA0562, CDCA3, STAG1, ANAPC1, ABCC10, NUDT11, CYP2E1, PARP10, CEP152, NCAPD3, NME7, CTDSPL2, PGM2, SPAG9, RFC3, TACSTD2, TANC2, TFPI, TUBA4A, TGFBRAP1, FAM177A1, AMFR, UNC13B, NUP43, GGCX, ALDOA, CRTC2, ORAI1, ENPP1, NEK2, PFKFB3, FAM122B, CACNB2, IGF2BP2, CHEK2, DIDO1, KIAA1009, SLC29A1, VRK1, FAM102A, NCAPG2, FBXO5, SLC4A1AP, WDHD1, NRBF2, RASEF, MAP2K4, NUF2, ATP11B, ZBTB40, CDC20, S100A14, HN1L, CKMT1A, IFI27, ATP13A1, HMGCS2, ARF1, C1ORF63, CD59, MAP7, SYTL2, FAM91A1, STEAP3, RTN4, CKMT1B, AP1B1, PPARG, ZNF295, DAXX, RTN3, CCNE2, ATAD3A, DNAJC15, AASDHPPT, HSF2, MAPT, PWWP2A, PARG, DOCK10, ERRFI1, TOP2A, DHCR24, GHR, CCNH, PI4KA, KIAA1430, SMN2, TNFRSF10A, SENP3, WDR47, H6PD, CKAP2L, RRM2, CCNYL1, TMSB4X, WDR44, USP24, THAP7, C1ORF93, SHROOM3, GULP1, SCML2, PPFIA1, UGDH, HK2, LARP1B, BCCIP, ZNF365, ZBTB16, GNG12, HMMR, LPXN, BEST1, NPAT, BCL6, BUB3, AGL, DTNA, POLR3G, SHMT1, NBPF10, DLGAP5, PAK1IP1, YWHAB, AFF3, BIRC5, CDKN3, FZD6, SDHA, COG3, ACCN2, ACSM3, AP2A2, PHF19, SFPQ, RASSF1, GFPT1, MAPK14, NARG2, KIF20B, ABCC4, LRP8, UBXN8, FCGR2A, WDR20, BCAR3, MRPS36, MTSS1, CEP72, ZC3HAV1, PRC1, PASK, EIF5B, PKMYT1, CSPG5, PI4K2B, ACSS2, TSSC1, SLC7A5, RELL1, COX4NB, ANKRD57, MLL5, KRT80, ELOVL5, OIP5, AAK1, STK39, ASPH, ASPM, TOR1AIP2, C2CD2, CARS, MADD, ENC1, SGOL1, TNFRSF14, TACC3, BICD2, TBC1D22A, TACC1, ELL2, MRTO4, FBXO16, KIAA1715, SLC38A1, CCDC18, AFTPH, ACTR3B, DIAPH1, DIAPH2, PNPT1, TIPIN, BMPR2, NOSTRIN, ANLN, FTH1, ATF2, ZFC3H1, POLE2, SLC30A1, HNRNPF, KATNA1, ORC6L, MNS1, RNF169, NDRG1, GNMT, SLC30A3, FBXO42, EIF3J, ASF1B, ATRIP, SLC30A6, DIXDC1, CARD9, EPB41, ATAD2, MALT1, MTRF1L, REEP1, KCTD5, REEP4, FOXP1, ATP7A, PDZD8, PLEKHA7, RFX1, WDR4, CPNE1, ABCC6P2, GPATCH4, CD79A, PIP4K2B, KIF23, MAD1L1, SEC24B, NBN, METAP2, C7ORF28B, SGPP1, E2F6, ALG3, ARHGAP19, HLCS, ACOX3, NFATC2IP, KIF2C, SLC1A5, MCTP2, AKR1C4, TMEM57, SLK, RNF103, PAK2, GRWD1, ITPRIP, URB1, RHOB, SNTB2, SHC1, C16ORF42, SHC2, PIGA, PRKCA, NOL6, ELP4, SGK3, PKN2, NUSAP1, RB1, TBCEL, PBK, MCM4, LMBRD2, VASP, MCM6, EYA3, ADRB2, EIF4G3, RIF1, ITGB1BP1, SNRNP200, TXNRD1, SPATS2L, HAUS8, LCP1, MELK, FKBP5, SSFA2, VIM, RDX, NFYA, DNAH2, CEP55, STUB1, PAIP2B, IARS, MTM1, ANXA9, ZNF512B, HJURP, MAP3K2, CENPA, TMEM154, SLC39A8, TUBA3D, TMEM87B, MSI2, GPNMB, LACTB, SLC39A3, TERF2, MOBKL3, CNNM4, TMEM45B, RAD51AP1, NCDN, NF1, PTPRA, C1ORF21, CENPF, CENPE, TMBIM1, RGS17, BAD, CDC25C, RACGAP1, MLF1IP, PTPN20A, MEMO1, SIRT2, CDC25B, ATXN1, IFIT2, ICK, DR1, 3.69E‐15 CENPV, FAF1, IGFBP3, SETD5 KIF23, MAD1L1, KIFC1, PRC1, NEK2, TIPIN, ANLN, PTTG1, CEP55, AURKB, KIAA1009, CCNE2, KIF2C, RNF103, NCAPG2, OIP5, INCENP, KATNA1, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, SGOL1, NUF2, CENPF, NUSAP1, CDC20, BIRC5, 4.52E‐15 CENPE, RACGAP1, CDC25C, SIRT2, TACC1, NCAPD3, CDC25B, CENPV, KIF20B, HAUS8, NUP43 KIF23, ADCY3, KIFC1, MAD1L1, NBN, PRC1, E2F6, PKMYT1, OSGIN2, AURKA, PTTG1, AURKB, RHOU, KIF2C, OIP5, INCENP, ASPM, CDCA3, STAG1, ANAPC1, SGOL1, MND1, NUSAP1, RB1, PBK, TACC3, NCAPD3, HAUS8, NUP43, NEK2, TIPIN, ANLN, CEP55, KIAA1009, NCAPG2, KATNA1, NPAT, MNS1, FBXO5, ZWILCH, BUB3, DLGAP5, NUF2, CENPF, BIRC5, CENPE, CDC20, 8.39E‐15 CDKN3, CDC25C, RAD54L, SIRT2, CDC25B, KIF20B, CENPV KIF23, MAD1L1, KIFC1, PRC1, NEK2, DIAPH2, TIPIN, ANLN, PTTG1, CEP55, ZBTB16, AURKB, DAXX, KIAA1009, CCNE2, RNF103, NCAPG2, OIP5, INCENP, KATNA1, FBXO5, ZWILCH, ASPM, STAG1, CDCA3, ANAPC1, SGOL1, NUF2, CENPF, NUSAP1, 3.51E‐14 CDC20, BIRC5, CENPE, RB1, RACGAP1, CDC25C, SIRT2, TACC1, NCAPD3, CDC25B, CENPV, KIF20B, HAUS8, NUP43 KIF23, MAD1L1, KIFC1, PRC1, NEK2, E2F6, TIPIN, PKMYT1, ANLN, AURKA, PTTG1, CEP55, AURKB, RHOU, KIAA1009, KIF2C, OIP5, NCAPG2, CENPA, INCENP, KATNA1, NPAT, FBXO5, ZWILCH, BUB3, ASPM, STAG1, CDCA3, ANAPC1, DLGAP5, SGOL1, NUF2, CENPF, NUSAP1, CDC20, CENPE, BIRC5, RB1, PBK, CDKN3, CDC25C, NCAPD3, SIRT2, CDC25B, CENPV, KIF20B, HAUS8, 4.05E‐13 NUP43
61 8.111702 72 9.574468
KIF23, ADCY3, MAD1L1, KIFC1, CEP72, NBN, PRC1, E2F6, PKMYT1, OSGIN2, AURKA, PTTG1, AURKB, DAXX, RHOU, KIF2C, MLL5, OIP5, INCENP, ASPM, DHCR24, CDCA3, STAG1, ANAPC1, SGOL1, MND1, NUSAP1, RB1, PBK, TACC3, NCAPD3, HAUS8, NUP43, NEK2, TIPIN, ANLN, CEP55, KIAA1009, NCAPG2, CENPA, KATNA1, NPAT, MNS1, FBXO5, ZWILCH, BUB3, DLGAP5, 6.06E‐13 NUF2, CENPF, BIRC5, CENPE, CDC20, RACGAP1, CDKN3, CDC25C, RAD54L, SIRT2, CDC25B, RASSF1, KIF20B, CENPV 5.56E‐12 ADCY3, STEAP3, KIF23, MAD1L1, KIFC1, CEP72, NBN, PRC1, E2F6, PKMYT1, OSGIN2, AURKA, AURKB, PTTG1, DAXX, RHOU,
SP_PIR_KEYWORDS phosphoprotein
SP_PIR_KEYWORDS cell division
GOTERM_BP_FAT GOTERM_BP_FAT
GO:0022402~cell cycle process GO:0007049~cell cycle
368 48.93617
42 5.585106
Table S3:GO terms that are enriched by HOTAIR‐repressed genes in LNCaP, Related to Figure 6. Category Term Count % PValue Genes LTBP4, LOC642590, MXI1, SOHLH2, AGXT2L2, N4BP2L2, SIN3B, DDX17, APP, ATP2B4, PICALM, KIAA1267, ZNF773, OGT, SCD5, BCL7A, BCL7C, C15ORF57, TMEFF2, RREB1, NOXA1, TMEM132A, DEPDC1, PTPRU, PIK3IP1, VASH2, SIRPA, ERGIC1, GRB10, SSTR2, UHRF2, RFC1, TBRG1, HES2, ZNF384, RALYL, PALM, C11ORF63, C3ORF15, MYO9A, ZNF618, FAM65B, SFRS15, SLC29A2, C19ORF29, SLC28A1, KIAA1683, SFRS12, IKZF2, PTGR2, FOXRED2, EPM2A, UAP1L1, ACACB, COL16A1, SNORD52, FNIP2, APOL3, SEMA6A, RNF7, SYTL1, NCOR2, RERE, DPP3, SH3RF1, ABCF3, HTATIP2, NRP1, DNAJC10, ZBBX, RTN1, MCCC2, AP2B1, NISCH, RRP1B, AEN, MYB, ANGPT2, LMOD3, ADCK4, FBXL17, SIK3, KIAA1949, ZBTB20, SF1, FLNB, C1QTNF9B, BNIP1, TAF15, THOC6, ZSCAN18, CYTSB, PARVA, CLUAP1, SHROOM4, TRIM13, NHLRC2, GCH1, MTMR3, COL7A1, CRISPLD2, C21ORF57, FBXW11, C21ORF58, SPATA7, TBXAS1, BRD3, IL8, NUCKS1, MEX3B, FADS2, KIAA1967, TBX1, PHF19, PHF14, FBLN2, MEX3D, MTSS1, PLXNA4, SLC44A4, ARNT2, IQGAP2, FAM179B, RGL1, KLHL7, GABPB1, SEZ6L2, ZNF187, RLN1, ST3GAL4, RLN2, SLC4A2, ITPK1, RMND1, INADL, C6ORF48, BEND4, BAIAP2, SLC22A23, MYO3B, CCDC74B, CECR5, TESK2, FAM86A, PTK7, LRIG1, ATHL1, ASTN2, RRAGB, SRC, CSMD1, PRR3, XAGE1C, SNF8, BTF3, AUTS2, RNF10, TMEM192, NDRG2, RAD51L3, TRAF4, SEC61A2, NIN, TAF5, GINS4, CD276, SACS, HACE1, EFS, SHANK3, FBXO38, NXN, FCAR, CACNA1H, CYFIP1, GRB7, ST5, NDST1, DEDD, NUP188, FAM63A, SDCBP2, ZNF816A, KLHL28, MKL2, CCDC88C, ARHGAP28, PCDHGB6, DGAT2, PDE9A, PMEPA1, PEG3, GALNT3, NUP98, ING3, ZC4H2, CYTH1, DNAH1, CYTH2, CALML4, KIAA0408, PRDM10, TNFRSF19, GTF3C1, PIK3R1, CHD3, MAF, TCIRG1, GSTA2, SLC12A2, CREB1, ATXN7L3, SIRT2, COL4A5, CCDC112, SP_PIR_KEYWORDS alternative splicing 220 45.36082 4.75E‐05 SON, INVS, ID1, BAX, C6ORF170, NFIB UP_SEQ_FEATURE domain:IQ 1 7 1.443299 7.92E‐05 MYO10, INVS, MYO3B, IQGAP3, IQGAP2, MYO9A, KIAA1683 UP_SEQ_FEATURE domain:IQ 2 7 1.443299 7.92E‐05 MYO10, INVS, MYO3B, IQGAP3, IQGAP2, MYO9A, KIAA1683 LTBP4, LOC642590, MXI1, SOHLH2, AGXT2L2, N4BP2L2, SIN3B, DDX17, APP, ATP2B4, PICALM, KIAA1267, ZNF773, OGT, SCD5, BCL7A, BCL7C, C15ORF57, TMEFF2, RREB1, NOXA1, TMEM132A, DEPDC1, PTPRU, PIK3IP1, VASH2, SIRPA, ERGIC1, SSTR2, GRB10, UHRF2, RFC1, TBRG1, HES2, ZNF384, RALYL, PALM, C11ORF63, C3ORF15, MYO9A, ZNF618, FAM65B, SFRS15, SLC29A2, C19ORF29, SLC28A1, KIAA1683, SFRS12, IKZF2, PTGR2, FOXRED2, EPM2A, UAP1L1, ACACB, COL16A1, SNORD52, FNIP2, APOL3, SEMA6A, RNF7, SYTL1, NCOR2, RERE, DPP3, SH3RF1, ABCF3, HTATIP2, NRP1, DNAJC10, ZBBX, RTN1, MCCC2, AP2B1, NISCH, RRP1B, AEN, MYB, ANGPT2, LMOD3, ADCK4, FBXL17, SIK3, KIAA1949, ZBTB20, SF1, FLNB, C1QTNF9B, BNIP1, TAF15, THOC6, CYTSB, ZSCAN18, PARVA, CLUAP1, SHROOM4, TRIM13, NHLRC2, GCH1, MTMR3, COL7A1, CRISPLD2, C21ORF57, FBXW11, C21ORF58, SPATA7, BRD3, IL8, NUCKS1, MEX3B, FADS2, KIAA1967, TBX1, PHF19, PHF14, MEX3D, MTSS1, PLXNA4, SLC44A4, ARNT2, IQGAP2, FAM179B, RGL1, KLHL7, GABPB1, SEZ6L2, ZNF187, RLN1, ST3GAL4, RLN2, SLC4A2, ITPK1, RMND1, INADL, C6ORF48, BEND4, BAIAP2, SLC22A23, MYO3B, CCDC74B, CECR5, TESK2, FAM86A, PTK7, LRIG1, ATHL1, ASTN2, RRAGB, SRC, CSMD1, PRR3, XAGE1C, SNF8, BTF3, AUTS2, RNF10, TMEM192, NDRG2, RAD51L3, TRAF4, SEC61A2, NIN, TAF5, GINS4, CD276, SACS, HACE1, EFS, SHANK3, FBXO38, NXN, FCAR, CACNA1H, CYFIP1, GRB7, ST5, NDST1, DEDD, NUP188, FAM63A, SDCBP2, ZNF816A, KLHL28, MKL2, CCDC88C, ARHGAP28, PCDHGB6, DGAT2, PDE9A, PMEPA1, PEG3, GALNT3, NUP98, ING3, ZC4H2, CYTH1, DNAH1, CYTH2, CALML4, KIAA0408, PRDM10, TNFRSF19, GTF3C1, PIK3R1, CHD3, MAF, TCIRG1, SLC12A2, CREB1, ATXN7L3, SIRT2, COL4A5, CCDC112, SON, INVS, ID1, BAX, UP_SEQ_FEATURE splice variant 217 44.74227 1.49E‐04 C6ORF170, NFIB zinc finger region:C2H2‐ ZNF84, ZNF559, RREB1, ZNF558, ZNF812, ZNF791, ZFP112, LOC389458, ZNF37A, ZNF837, ZNF304, ZNF187, ZNF773 UP_SEQ_FEATURE type 8 24 4.948454 1.86E‐04 ZNF816A, PRDM10, ZNF384, ZNF286A, ZNF416, ZNF579, ZNF426, ZNF471, ZNF432, ZNF571, PEG3 ZNF84, ZNF559, ZNF558, ZFP112, ZNF304, ZNF187, ZNF816A, ZNF773, PRDM10, ZNF286A, ZNF579, ZNF426, ZNF471 zinc finger region:C2H2‐ type 7 25 5.154639 2.30E‐04 RREB1, ZNF812, ZNF791, LOC389458, ZNF37A, ZNF837, SALL2, ZNF384, ZNF416, ZNF432, ZNF571, PEG3 UP_SEQ_FEATURE zinc finger region:C2H2‐ ZNF84, ZNF559, ZNF558, ZFP112, ZNF304, ZNF187, ZNF816A, ZNF773, PRDM10, ZNF286A, ZNF579, ZNF426, ZNF471, IKZF2 UP_SEQ_FEATURE type 6 26 5.360825 3.07E‐04 RREB1, ZNF812, ZNF791, LOC389458, ZNF37A, ZNF837, SALL2, ZNF384, ZNF416, ZNF432, ZNF571, PEG3 ZNF84, SH3RF1, ZBTB34, UTRN, RNF215, ZBBX, RNF214, ZNF304, ZNF187, ZNF816A, ZNF773, ZNF579, ZBTB20, RREB1 ZNF812, SF1, ZHX2, ZNF791, MORC2, ZNF37A, UHRF2, EP300, TAF15, ZNF384, ZSCAN18, ZNF432, PEG3, ING3, ZNF559, ZNF558, ZC4H2, TRIM13, ZFP112, MYO9A, ZNF618, NR2C2, TRIM65, MTMR3, PRR3, PRDM10, ZNF286A, ZNF609, ZNF471, ZNF426, RNF10, TRAF4, CHD3, RNF144A, IKZF2, MEX3B, LOC389458, ATXN7L3, ZNF837, MNAT1, SALL2, RNF44, PHF2, SP_PIR_KEYWORDS zinc‐finger 64 13.19588 3.50E‐04 RNF7, PHF19, PHF14, ZNF416, MEX3D, ZNF571, RERE DPP3, SH3RF1, ZNF84, ZBTB34, UTRN, ADH1C, ADH1A, RNF215, ZBBX, RNF214, ZNF304, APP, ZNF187, ZNF816A, ZNF773 TIMM9, ZNF579, ZBTB20, RREB1, ZNF812, SF1, ZHX2, ZNF791, MORC2, ZNF37A, MMP11, EP300, UHRF2, TAF15, ZNF384, PDE9A, ZNF432, ZSCAN18, PEG3, ING3, ZNF559, ZC4H2, ZNF558, TRIM13, ZFP112, ZNF618, MYO9A, NR2C2, GCH1, TRIM65, MTMR3, PRR3, RPS29, PRDM10, SLC30A4, ZNF286A, C21ORF57, SLC39A7, ZNF609, ZNF471, ZNF426, RNF10, TRAF4, CHD3, RNF144A, IKZF2, MEX3B, MOBKL1A, LOC389458, SIRT2, ATXN7L3, ZNF837, MNAT1, SALL2, RNF44, PHF19, PHF2, RNF7, SP_PIR_KEYWORDS zinc 77 15.87629 4.37E‐04 PHF14, ZNF416, MEX3D, ZNF571, RERE
SP_PIR_KEYWORDS
phosphoprotein
SP_PIR_KEYWORDS
dna‐binding
SP_PIR_KEYWORDS
nucleus zinc finger region:C2H2‐ type 12 domain:IQ 3
UP_SEQ_FEATURE UP_SEQ_FEATURE
208
42.8866
67 13.81443
131 27.01031 16 3.298969 5 1.030928
UTRN, LOC642590, RNF215, RNF214, N4BP2L2, DDX17, APP, ATP2B4, KIAA1267, MC1R, OGT, BCL7A, BCL7C, KRR1, RREB1, NOXA1, ZHX2, DEPDC1, PTPRU, SIRPA, SSTR2, GRB10, UHRF2, AAAS, F5, RFC1, ZNF384, TGIF2, CCDC97, CRTC2, PALM, LMNB1, C3ORF15, MYO9A, EPHB4, ZNF618, FAM65B, SFRS15, SLC29A2, RAC3, C19ORF29, ALDH4A1, CDC42EP3, GCNT1, KIAA1683, SFRS12, ST6GAL1, IKZF2, ACACB, FNIP2, RNF7, MCPH1, POP1, SYTL1, NCOR2, RERE, RANBP10, DPP3, SH3RF1, ABCF3, NRP1, HTATIP2, RTN1, MTHFD1, AP2B1, NISCH, RRP1B, DYNC1H1, JPH1, TUBB3, SIK3, USP14, TUBB4, KIAA1949, ZBTB20, SF1, FLNB, MORC2, ST6GALNAC1, TNFRSF10A, RAB11FIP2, TAF15, HIST2H2BF, CYTSB, FKBP10, CLOCK, PARVA, CLUAP1, SHROOM2, HIST1H2AE, TRIM13, NHLRC2, GCH1, MTMR3, FAT1, CD24, HIST1H2BC, BRD3, NUCKS1, MEX3B, HIST1H2BG, KIAA1967, MAPK11, SF3A1, RPL28, MNAT1, RPS6KA3, C12ORF35, PHF19, PSMD13, PHF14, FBLN2, CSNK1E, MEX3D, MTSS1, PLXNA3, PLXNA4, F2RL1, IQGAP3, IQGAP2, FAM179B, INO80, PRKCSH, RGL1, SH2D5, LOC643778, DNAJC7, ZNF579, SLC4A2, INADL, BEND4, BAIAP2, GAGE12F, GAGE12E, GAGE12B, EP300, TELO2, TESK2, FAM171A1, STK16, SOX4, ITGB2, TTC5, RIOK1, LLGL1, NR2C2, SRC, SBK1, BLOC1S1, ADRA2A, BTF3, AUTS2, TMEM192, NDRG2, KIAA1211, AXIN2, TMEM79, TRAF4, PTPN9, NIN, TBC1D10A, FSCN1, CD276, SACS, MOBKL1A, EFS, SLC6A16, SHANK3, FBXO38, SALL2, MYO10 PHF2, FCAR, CACNA1H, CYFIP1, AHCYL2, GRB7, ST5, ARHGAP10, NUP188, FAM63A, SPRY1, MKL2, CCDC88C, ARHGAP28, NCOA5, C5ORF13, GAA, PDE9A, NUP98, RNH1, DNAH1, PRDM10, ENO2, SLC39A7, GEMIN8, ZNF609, GTF3C1, ERCC4, 5.13E‐04 PIK3R1, CHD3, SLC12A2, CREB1, DPYSL2, COTL1, ATXN7L3, SIRT2, PHAX, SON, ID1, NFIB, F2R ZNF84, ZBTB34, DEDD, TBX21, ARNT2, INO80, MXI1, SOHLH2, ZNF304, DDX12, ZNF816A, ZNF773, ZNF579, MYB, TWIST1 ZBTB20, RREB1, ZNF812, ZHX2, ZNF791, MXD1, TOX3, ZNF37A, UHRF2, TAF15, RFC1, HIST2H2BF, HES2, ZNF384, TGIF2, ZNF432, ZSCAN18, CLOCK, PEG3, ZNF559, ZNF558, HIST1H2AE, SOX4, SOX9, ZFP112, SOX8, ZNF618, NR2C2, HMGXB4, PRDM10, ZNF286A, ZNF471, ZNF426, RAD51L3, GTF3C1, ERCC4, CHD3, MAF, MAFF, HIST1H2BC, IKZF2, HIST1H2BG, CREB1, 6.87E‐04 TEAD2, TBX1, ZNF837, PKNOX2, SALL2, SON, ZNF416, ZNF571, NCOR2, NFIB PNMA2, ARNT2, INO80, MXI1, SOHLH2, KLHL7, ZNF304, GABPB1, DDX17, SIN3B, ZNF187, DDX12, ZNF773, ZNF579, OGT, TWIST1, KRR1, RREB1, ZNF812, ZHX2, ZNF791, DEPDC1, TOX3, ZNF37A, EP300, UHRF2, RFC1, TELO2, HES2, TBRG1, ZNF384, TESK2, TGIF2, ZNF432, CRTC2, CDK5R1, LMNB1, SOX4, TTC5, SOX9, SOX8, ZNF618, NR2C2, SFRS15, SLC29A2, SNF8, BTF3, ZNF426, RAD51L3, TRAF4, SFRS12, MAFF, IKZF2, NOC4L, TAF5, GINS4, EPM2A, MOBKL1A, TEAD2, ZNF837, FBXO38, SALL2, RNF7, PHF2, NXN, POP1, ZNF416, NCOR2, RERE, HINT3, ZNF84, ZBTB34, HTATIP2, DEDD, TBX21, NUP188, RRP1B, ZNF816A, AEN, MKL2, MYB, ZBTB20, SF1, MXD1, TAF15, HIST2H2BF, NCOA5, CYTSB, ZSCAN18, CLOCK, PEG3, CLUAP1, YPEL1, ING3, NUP98, ZNF559, ZNF558, HIST1H2AE, ZFP112, GCH1, HMGXB4, PRDM10, ZNF286A, ZNF471, ZNF609, GEMIN8, GTF3C1, ERCC4, CHD3, MAF, HIST1H2BC, BRD3, NUCKS1, MEX3B, HIST1H2BG, CREB1, TBX1, LOC389458, SF3A1, ATXN7L3, PKNOX2, 0.001041 PHAX, MNAT1, SON, PHF19, ID2, INVS, ID1, DET1, MEX3D, ZNF571, NFIB ZNF84, ZNF559, RREB1, ZNF812, ZNF791, LOC389458, ZFP112, ZNF37A, ZNF304, ZNF816A, ZNF416, ZNF426, ZNF471 0.001119 ZNF432, ZNF571, PEG3 0.001144 MYO10, IQGAP3, IQGAP2, MYO9A, KIAA1683
Table S4. oligonucleotide sequences used in this study, Related to Figure 1. Primer Names HOTAIR F1 HOTAIR R1 U1 F1 U1 R1 AR F1 AR R1 PSA F1 PSA R1 KLK2 F1 KLK2 R1 TMPRSS2 F1 TMRPSS2 R1 NOV F2 NOV R2 OPRK1 F1 OPRK1 R1 FKBP5 F1 FKBP5 R1 NKX3.1 F1 NKX3.1 R1 DDC F1 DDC R1 SI F1 SI R1 MET F1 MET R1 KIAA0066 pF1 KIAA0066 pR1 PSA pF1 PSA pR1 TMPRSS2_pF1 TMPRSS2_pR1 FKBP5 pF1 FKBP5 pR1 OPRK1 pF1 OPRK1 pR1 MET pF1 MET pR1 HOTAIR pF1 HOTAIR pR1 Hotair p F2 Hotair p R2 Hotair P F3 Hotair P R3 Hotair H5 F1 Hotair H5 R1
Sequence GGCGGATGCAAGTTAATAAAAC TACGCCTGAGTGTTCACGAG ATACTTACCTGGCAGGGGAG CAGGGGGAAAGCGCGAACGCA CAGTGGATGGGCTGAAAAAT GGAGCTTGGTGAGCTGGTAG ACGCTGGACAGGGGGCAAAAG GGGCAGGGCACATGGTTCACT CCATGCCTGGAGACATATCA TCCAGCACATGTCACTCTCC CAGGAGTGTACGGGAATGTGATGGT GATTAGCCGTCTGCCCTCATTTGT ACCGTCAATGTGAGATGCTG TCTTGAACTGCAGGTGGATG AACTCGCTGGTCATGTTCGT CTCTGAAAGGGCATGGTTGT TCTCATGTCTCCCCAGTTCC TTCTGGCTTTCACGTCTGTG GAGACGCTGGCAGAGACC CACCTGAGTGTGGGAGAAGG GCCCCTACTTCTTCGCCTAC CACAGTCTCCAGCTCTGTGC TTTTGGCAGCCTTATCCAAG CAATCAGAGAGATTTCCAATCCA CGCTGACTTCTCCACTGGTT TACACTCCCCATTGCTCCTC CTAGGAGGGTGGAGGTAGGG GCCCCAAACAGGAGTAATGA GCCTGGATCTGAGAGAGATATCATC ACACCTTTTTTTTTCTGGATTGTTG GGTAAACTCTCCCTGCCACA TACTCCAGGAAGTGGGGATG GGTTCCTGGGCAGGAGTAAG AACGTGGATCCCACACTCTC CTGCTCCTGGCATTATCCTC TGTGGCTCTCAGCAGGAAGT GCTGCTTTGGGAAATGGTTA TGGTCGCAGGTTTCAACATA AGCTGTTTGTTCCCTTTTGC TCGCGGCATTTTTATGAGAT TGGCCTGTTTATATCCCTTCC CCCAAAACCATTTCCTGAGA AGCACCTCCGGGATATTAGG AGAGAGCACCAGGCACTGAG AGCCTTTGGAAGCTCTTGAA GTGTCTTGGAGAGGCGTGTA
Hotair T7‐5.1
TAATACGACTCACTATAGGGACATTCTGCCCTGATTTCCG
Hotair Not I 3.2
GCGGCCGCAAAGACGCCCCTCCTTCCTC
Hotair T7‐5.1
TAATACGACTCACTATAGGGACATTCTGCCCTGATTTCCG
Hotair NotI‐3.4
GCGGCCGC TCACTTTTAAAAATTTGTTT
HotairT7‐5.3
TAATACGACTCACTATAGGGAGTTCCCGGAACAAACGTGG
Hotair NotI‐3.3
GCGGCCGC GAAAATGCATCCAGATATTA
Hotair T7‐5.2
TAATACGACTCACTATAGGGAACCAGCCCTAGCCTTTGGA
Hotair NotI 3.3 GCGGCCGC GAAAATGCATCCAGATATTA Hot NotI 5.1(anti)
GCGGCCGCACATTCTGCCCTGATTTCCG
Hot T7‐3.1 (anti) Hotair‐R1‐5.1 Hotair‐Not1‐3.1 Hotair‐ARE1‐m‐5.1 Hotair‐ARE1‐m‐3.1 Hotair‐ARE2‐m‐5.1 Hotair‐ARE2‐m‐3.1 AR‐BamH1‐5.1 AR‐NT‐Flag‐Xho1‐3.1 AR‐DBD‐Flag‐Xho1‐3.1 AR‐DBD‐BamH1‐5.1 AR‐LBD‐Flag‐Xho1‐3.1 AR‐LBD‐BamH1‐5.1 siHOTAIR #1 siHOTAIR #2
TAATACGACTCACTATAGGGAAAGACGCCCCTCCTTCCTC ATTCTCTAGAATTCACATTCTGCCCTGATTTCCG ATTCTCTAGGCGCGCCGAAAATGCATCCAGATATTA GTAAGAGAGCACCAGGCATTGCGTCCTGAATGTGAAACAGACCAACACCCCTGC GCAGGGGTGTTGGTCTGTTTCACATTCAGGACGCAATGCCTGGTGCTCTCTTAC CGGCTCCCACCCGGGACAAGGACTTACAGTAACCGAAAATCCCGGAGGT ACCTCCGGGATTTTCGGTTACTGTAAGTCCTTGTCCCGGGTGGGAGCCG GAGCTCGGATCCCAGCCAAGCTCAAGGATGGA TCTAGACTCGAGTCACTTATCGTCGTCATCCTTGTAATCCATTTCGCTTTTGACACAAG TCTAGACTCGAGTCACTTATCGTCGTCATCCTTGTAATCGGTGCTGGAAGCCTCTCCTT GAGCTCGGATCCGGGCCCACCATGGGCCCCTGGATGGATAGCTA TCTAGACTCGAGTCACTTATCGTCGTCATCCTTGTAATCCTGGGTGTGGAAATAGATGG GAGCTCGGATCCGGGCCCACCATGACCAGCCCCACTGAGGAGAC GAACGGGAGUACAGAGAGAUU CCACAUGAACGCCCAGAGAUU
Application qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR qRT‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR ChIP‐PCR clone HOTAIR 1‐360bp fragment for in vitro transcription clone HOTAIR 1‐360bp fragment for in vitro transcription clone HOTAIR 1‐1550 bp fragment for in vitro transcription clone HOTAIR 1‐1550 bp fragment for in vitro transcription clone HOTAIR 660‐ 2337bp fragment for in vitro transcription clone HOTAIR 660‐ 2337bp fragment for in vitro transcription clone HOTAIR 1550‐ 2337bp fragment for in vitro transcription clone HOTAIR 1550‐ 2337bp fragment for in vitro transcription clone antisense HOTAIR 1‐ 2337bp fragment for in vitro transcription clone antisense HOTAIR 1‐ 2337bp fragment for in vitro transcription clone HOTAIR ARE mutants clone HOTAIR ARE mutants clone HOTAIR ARE mutants clone HOTAIR ARE mutants clone HOTAIR ARE mutants clone HOTAIR ARE mutants clone AR deletion mutants clone AR deletion mutants clone AR deletion mutants clone AR deletion mutants clone AR deletion mutants clone AR deletion mutants clone shHOTAIR #1 clone shHOTAIR #2
