Supporting Information

Renaud et al. 10.1073/pnas.1507397112

Fig. S1. Gene pathway analysis comparing trophoblast cells cultured under undifferentiated or differentiating conditions. (A) Pie chart showing the number of transcripts with altered expression levels in BeWo trophoblast cells following 24-h exposure to differentiating (Diff) conditions compared with cells maintained in undifferentiating (Undiff) conditions. (B) DAVID pathway analysis of genes significantly altered following BeWo trophoblast differentiation.

Renaud et al. www.pnas.org/cgi/content/short/1507397112 1of9 Fig. S2. Expression of CGA and CGB in primary cytotrophoblast cells isolated from term placenta. Primary trophoblast cells were isolated and plated for 6-, 24-, or 72-h duration. RNA was extracted at each time point, and relative expression of CGA and CGB transcript was examined by qRT-PCR compared with 18S rRNA. *P < 0.05; n = 3.

Fig. S3. Expression of OVOL1 and OVOL2 in mouse and rat trophoblast cells. (A) RT-PCR analysis of mouse and rat placenta (Plac) and mouse and rat TS cells for Ovol1 and Ovol2. 18S rRNA was used as a loading control. Mouse and rat skin were used as positive controls (+). D, TS cells under differentiating conditions (8 d); M, DNA ladder; S, TS cells maintained in stem conditions. (B) Immunohistochemistry for OVOL2 (red) in mouse and rat placenta. Nuclei are counterstained blue using DAPI. Note the OVOL2+ trophoblast giant cells at the interface of the placenta and decidua (white arrows). Insets show a high-magnification view of the labyrinth zone. (Scale bar, 100 μm.)

Renaud et al. www.pnas.org/cgi/content/short/1507397112 2of9 Fig. S4. OVOL1 regulates syncytiotrophoblast gene expression in human iPS cells induced to differentiate along the trophoblast lineage. Human iPS cells were maintained in FGF2 (undifferentiated, Undiff) or BMP4 (differentiating, Diff) for 7 d. (A) Transcript expression of CGA, CGB, ERVW-1, ERVFRD-1, ERVV-1,andERVV-2 in human iPS cells maintained in undifferentiated or differentiating conditions. (B) CGB immunofluorescence in human iPS cells maintained in undifferentiated or differentiating conditions. (Scale bars, 100 μm.) (C and D)Transcript(C) and expression (D) of OVOL1 in human iPS cells maintained in undifferentiated or differentiating conditions. (E) Effect of OVOL1 deficiency (shOVOL1) in human iPS cells maintained in differentiating conditions on endogenous retrovirus-derived gene expression (ERVW-1, ERVFRD-1, ERVV-1,andERVV-2) compared with cells expressing control shRNA (shCTRL). *P < 0.05; n = 3.

Fig. S5. Effect of OVOL1 knockdown in primary cytotrophoblast cells on genes linked with the cytotrophoblast progenitor state. Transcript expression of , ID1, TP63, and ASCL2 in primary cytotrophoblasts transduced with shRNAs targeting OVOL1 (shOVOL1). *P < 0.05; n = 6.

Renaud et al. www.pnas.org/cgi/content/short/1507397112 3of9 Fig. S6. OVOL1 was not detected binding genomic regions near ERVFRD-1 and did not affect GCM1 expression. (A) ChIP was conducted in lysates of BeWo trophoblast cells maintained in differentiating conditions using antibodies for OVOL1 or IgG (negative control). Primers were designed to span regions containing putative OVOL1-binding sites (indicated by inverted triangles) near ERVFRD-1.(B) Nuclear protein expression of GCM1 in trophoblast cells cultured under differentiating conditions for 6, 24, or 48 h. Cells maintained in undifferentiated conditions (Undiff; 0 h) were used as controls. (C and D) BeWo tro- phoblast cells expressing control shRNA (shCTRL) or shRNAs targeting OVOL1 (shOVOL1-a or shOVOL1-b) were maintained under undifferentiated or dif- ferentiating (Diff) conditions for 48 h. Subsequently, qRT-PCR (C) and Western blotting (D) were conducted to detect transcript and nuclear protein of GCM1, respectively. TFIID was used as a loading control for Western blotting. *P < 0.05, OVOL1-deficient trophoblast cells vs. shCTRL-expressing trophoblast cells; n = 3; #P < 0.05, trophoblast cells maintained in undifferentiated vs. differentiating conditions; n = 3.

Renaud et al. www.pnas.org/cgi/content/short/1507397112 4of9 Table S1. Top up-regulated and down-regulated transcripts following BeWo trophoblast differentiation Gene Undifferentiated Differentiating FoldΔ Gene name

Up-regulated transcripts PRR9 136.31 1,482.43 10.88 Proline rich 9 CRIP2 65.28 527.13 8.07 Cysteine-rich protein 2 GREB1 125.15 895.39 7.15 GREB1 protein MMP12 48.45 344.33 7.11 Matrix metallopeptidase 12 (macrophage elastase) TGM2 73.62 522.15 7.09 Transglutaminase 2 OVOL1 1,445.95 8607.53 5.95 Ovo-like 1(Drosophila) CGB 3,698.46 2,1043.60 5.69 CG; β polypeptide SLC6A19 540.41 3052.59 5.65 Solute carrier family 6 (neutral amino acid transporter); member 19 LHB 683.57 3674.48 5.38 Luteinizing hormone beta polypeptide SLC19A1 67.21 351.21 5.23 Solute carrier family 19 (folate transporter); member 1 CGA 469.15 2,440.61 5.20 Glycoprotein hormones; alpha polypeptide OVOL1 199.01 1,016.53 5.11 Ovo-like 1(Drosophila) TFF1 729.03 3,712.17 5.09 Trefoil factor 1 CORO1A 138.75 701.04 5.05 Coronin; actin-binding protein; 1A INSL4 139.38 692.86 4.97 Insulin-like 4 (placenta) COL27A1 96.62 446.13 4.62 Collagen; type XXVII; alpha 1 SERPINE1 825.81 3,636.59 4.40 Serpin peptidase inhibitor; clade E, member 1 RSAD2 162.93 713.58 4.38 Radical S-adenosyl methionine domain containing 2 SIK1 628.01 2,664.13 4.24 Salt-inducible kinase 1 PTPN21 312.58 1,298.91 4.16 Protein tyrosine phosphatase; nonreceptor type 21 FOSL2 74.05 302.84 4.09 FOS-like antigen 2 SGK1 1,308.83 5,322.08 4.07 Serum/glucocorticoid regulated kinase 1 HSD3B1 183.95 744.51 4.05 Hydroxy-δ-5-steroid dehydrogenase; ANGPT4 150.97 607.60 4.02 Angiopoietin 4 WIPF1 217.95 852.72 3.91 WAS/WASL interacting protein family; member 1 CD59 813.51 3,146.33 3.87 CD59 molecule; complement regulatory protein CXCR7 1,446.51 5,479.55 3.79 Chemokine (C-X-C motif) 7 DUSP4 138.23 512.31 3.71 Dual specificity phosphatase 4 TGM2 124.98 460.57 3.69 Transglutaminase 2 RSAD2 132.11 483.21 3.66 Radical S-adenosyl methionine domain containing 2 VGLL3 93.74 341.83 3.65 Vestigial like 3 (Drosophila) CD59 930.67 3,371.21 3.62 CD59 molecule; complement regulatory protein MXD1 573.73 2,066.21 3.60 MAX dimerization protein 1 SP6 1,781.73 6,398.21 3.59 Sp6 FGFR3 591.76 2,105.17 3.56 Fibroblast growth factor receptor 3 HOPX 1,437.74 5,054.33 3.52 HOP PMAIP1 459.45 1,576.06 3.43 Phorbol-12-myristate-13-acetate-induced protein 1 NDRG1 368.75 1,255.84 3.41 N-myc downstream regulated 1 LAIR2 3,202.28 10,751.16 3.36 Leukocyte-associated Ig-like receptor 2 ADAM33 90.26 302.81 3.36 ADAM metallopeptidase domain 33 C15orf48 320.76 1,074.67 3.35 15 ORF 48 FAM43A 585.31 1,946.03 3.32 Family with sequence similarity 43; member A ST8SIA4 314.41 1,043.62 3.32 ST8 alpha-N-acetyl-neuraminide alpha-2;8-sialyltransferase 4 HERV-FRD 1,200.14 3,946.61 3.29 HERV-FRD provirus ancestral Env polyprotein Down-regulated transcripts ST3GAL5 825.20 166.31 0.20 ST3 beta-galactoside alpha-2;3-sialyltransferase 5 OVOL2 1,094.92 229.17 0.21 Ovo-like 2 (Drosophila) RNF144B 1,295.38 322.93 0.25 Ring finger protein 144B ENC1 1,766.98 441.29 0.25 Ectodermal-neural cortex (with BTB-like domain) LOC645431 539.58 142.95 0.26 Hypothetical LOC645431 SCARA3 478.81 127.07 0.27 Scavenger receptor class A; member 3 NANOS1 997.31 302.30 0.30 Nanos homolog 1 (Drosophila) IL17RD 512.59 160.32 0.31 Interleukin 17 receptor D NR2E1 327.66 103.21 0.31 subfamily 2; group E; member 1 RNF125 399.46 127.27 0.32 Ring finger protein 125 TP63 530.14 169.93 0.32 Tumor protein p63 MYOZ1 322.70 103.64 0.32 Myozenin 1 VTCN1 399.07 128.22 0.32 V-set domain containing T-cell activation inhibitor 1 KANK4 1,662.05 539.58 0.32 KN motif and ankyrin repeat domains 4 GRLF1 458.36 149.14 0.33 DNA-binding factor 1 TCF7L1 452.03 151.93 0.34 Transcription factor 7-like 1 (T-cell specific; HMG-box) DFNA5 1,995.57 688.43 0.34 Deafness; autosomal dominant 5

Renaud et al. www.pnas.org/cgi/content/short/1507397112 5of9 Table S1. Cont. Gene Undifferentiated Differentiating FoldΔ Gene name

ANGPTL2 467.28 161.72 0.35 Angiopoietin-like 2 WNT11 315.07 110.59 0.35 Wingless-type MMTV integration site family; member 11 NEXN 492.66 175.57 0.36 Nexilin (F actin-binding protein) FAM189A2 397.21 141.97 0.36 Family with sequence similarity 189; member A2 C14orf139 2,202.68 787.43 0.36 Chromosome 14 ORF 139 C1orf105 777.45 278.05 0.36 Chromosome 1 ORF 105 CBX5 649.78 233.09 0.36 Chromobox homolog 5 (HP1 alpha homolog; Drosophila) SORBS2 305.77 112.35 0.37 Sorbin and SH3 domain containing 2 HAND1 592.20 220.10 0.37 Heart and neural crest derivatives expressed 1 TRERF1 560.82 210.39 0.38 Transcriptional regulating factor 1 STAT1 1,706.06 643.62 0.38 Signal transducer and activator of transcription 1; 91kDa RHPN2 1,139.72 430.29 0.38 Rhophilin; Rho GTPase-binding protein 2 MOCS1 434.60 164.57 0.38 Molybdenum cofactor synthesis 1 LOC729680 3,849.00 1,457.97 0.38 Hypothetical protein LOC729680 TMEM139 588.01 224.15 0.38 Transmembrane protein 139 CRAT 3,014.61 1,153.44 0.38 Carnitine acetyltransferase KRCC1 1,270.83 489.98 0.39 Lysine-rich coiled-coil 1 ANXA9 315.26 122.06 0.39 Annexin A9 PIK3R3 1,846.28 726.41 0.39 Phosphoinositide-3-kinase; regulatory subunit 3 (gamma) KRCC1 601.78 237.99 0.40 Lysine-rich coiled-coil 1 RBPMS 2,294.36 910.26 0.40 RNA-binding protein with multiple splicing TAF12 997.12 395.76 0.40 TAF12 RNA polymerase II PMP22 1,678.82 675.47 0.40 Peripheral myelin protein 22 HP/HPR 348.54 141.90 0.41 Haptoglobin/haptoglobin-related protein HUNK 1,185.84 487.24 0.41 Hormonally up-regulated Neu-associated kinase FAM119A 989.39 406.67 0.41 Family with sequence similarity 119; member A FBXO11 370.82 152.53 0.41 F-box protein 11

OVOL1 is shown in boldface.

Renaud et al. www.pnas.org/cgi/content/short/1507397112 6of9 Table S2. Transcripts altered more than twofold following OVOL1 knockdown Gene shCTRL RPKM shOVOL1 RPKM FoldΔ Gene shCTRL RPKM shOVOL1 RPKM FoldΔ

PHOSPHO1 2.15 0.12 −9.72 AC007566.10 22.81 9.54 −2.54 PRR9 29.96 4.61 −6.81 HSPB8 9.41 3.92 −2.52 ERVFRD-1 70.47 11.96 −6.28 SDC1 15.14 6.38 −2.52 HSD11B2 15.61 3.07 −5.28 SQSTM1 26.14 11.19 −2.49 RP11-463P17.1 8.61 1.65 −5.25 RP11-770G2.5 133.23 57.62 −2.48 SMIM13 16.55 3.63 −4.77 C5orf45 20.30 8.75 −2.47 GADD45G 16.03 3.88 −4.33 RP11-1100L3.8 21.93 9.54 −2.45 TREML2 24.33 6.30 −4.09 CGB8 955.83 419.18 −2.45 HSPB1P2 11.12 2.83 −4.08 ALAS1 11.61 5.05 −2.43 RIN3 7.95 2.04 −4.00 FHDC1 11.44 4.98 −2.43 LGALS16 7.09 1.83 −3.94 OVOL1 32.45 14.31 −2.42 HSD3B1 3.08 0.75 −3.92 CGB1 234.75 105.09 −2.40 4.55 1.18 −3.84 RP5-943J3.2 7.03 3.09 −2.38 DUSP5 5.12 1.37 −3.75 PITX1 9.59 4.26 −2.38 HSPB1P1 14.63 4.19 −3.67 PMAIP1 15.98 7.17 −2.37 HSPB1 7.32 2.06 −3.65 RP11-106M3.1 10.59 4.88 −2.29 AL590714.1 16.98 4.90 −3.65 CGB5 551.48 258.54 −2.29 APOA2 27.30 7.97 −3.63 ALPPL2 12.81 5.96 −2.28 INSL4 3.82 1.07 −3.51 GDF15 34.25 16.49 −2.22 SLC7A8 7.96 2.38 −3.45 AC068134.8 6.88 3.30 −2.19 CGB7 15.33 4.69 −3.44 KATNBL1P6 22.07 10.90 −2.16 ERVV-1 162.23 50.94 −3.42 RP11-169L17.5 8.43 4.13 −2.15 CTD-2620I22.3 61.52 19.25 −3.42 RP11-809N8.2 19.62 9.88 −2.12 KRT23 6.50 1.98 −3.37 NUS1P1 99.28 50.48 −2.11 LYPD3 14.46 4.76 −3.20 EAF1 32.42 16.46 −2.10 CGB2 6.27 2.03 −3.18 EAF1-AS1 30.16 15.35 −2.10 JUNB 7.38 2.45 −3.12 RP11-457K10.2 33.08 16.87 −2.10 ERVV-2 47.46 16.39 −3.09 NUPR1 47.17 24.16 −2.09 SLC40A1 7.33 2.55 −2.98 FDX1P1 75.45 38.62 −2.09 FOXI3 6.22 2.17 −2.96 DNAJB9 16.25 8.36 −2.07 LEP 9.75 3.71 −2.76 RHOB 22.56 11.73 −2.05 ZNF702P 16.85 6.56 −2.72 C2orf72 9.35 4.83 −2.05 TBX3 15.39 6.00 −2.71 FBRS 7.72 3.98 −2.05 TNFRSF1B 6.39 2.45 −2.71 KRT8P33 12.59 6.63 −2.02 CCR7 39.72 15.74 −2.69 CTSL 25.58 13.54 −2.02 CGA 543.86 217.76 −2.69 AC007620.3 3.48 8.60 2.26 ERVW-1 77.97 31.46 −2.65 RP11-734J24.1 3.49 8.65 2.27 CTB-60B18.6 11.58 4.71 −2.59 RP11-145M9.4 3.87 10.14 2.40 ARL4C 11.79 4.87 −2.56 RP11-349N19.2 17.63 46.61 2.44 CGB 231.88 97.77 −2.55

Renaud et al. www.pnas.org/cgi/content/short/1507397112 7of9 Table S3. Primers used for RT-PCR Gene Accession no. Forward primer Reverse primer Size, bp

Primers used for qRT-PCR RNA18S5 NR_003286.2 GCAGCCCAAACTAACCTCAC TAGCCATAAGGTCCGCTCTC 123 APOA2 NM_001643.1 TTCGTGGAACTTGGAACACA GCAAAGAGTGGGTAGGGACA 124 ASCL2 NM_005170.2 AACTTGAGCTGCTGGAGGGACA TCTTGGCCAGCATGGAAAACTC 132 CCR7 NM_001838.3 GGAGACTTCTTGGCTTGGTG GTGAGGGGATGAGTGTGCTT 148 CGA NM_000735.3 TTTCTGGTCACATTGTCGGT TGGGCAATCCTGCACATCAG 65 CGB NM_000737.3 CCTGGCCTTGTCTACCTCTT GGCTTTATACCTCGGGGTTG 108 CORO1A NM_007074.3 TGCACCCAGACACGATCTAC CGGTCCTTCTCAGCTACGAC 126 CTD-2620I22.3 HG511000.1 CCACAAAAACCCAAGAGGAA AGGGCCTAGGAAAGTGGAGA 104 ENC1 NM_003633.3 CCGTGACCGTCGCTAGAA ACGCTGCTGGCGTAAAAG 192 ERVFRD-1 NM_207582.2 CCAAATTCCCTCCTCTCCTC CGGGTGTTAGTTTGCTTGGT 115 ERVH48-1 BC005107.1 AGGTAGGAGGGAGCCAAAGA AGTGCGATAACCCCATGAAC 109 ERVV-1 NM_152473.2 TAACAGTGGGGCGATAGAGG AGACTTCACAGCCTCCCAAA 111 ERVV-2 NM_001191055.1 CAGGCACAGTGGAATGAAAA GACCTGGTGATGAAGTTGTGG 103 ERVW-1 NM_014590.3 CTACCCCAACTGCGGTTAAA GGTTCCTTTGGCAGTATCCA 84 GCM1 NM_003643.3 GCAACACCAACAACCACAAC GTAAATCTTGCGGCCTTCCT 100 HSD11B2 NM_000196.3 CAGATGGACCTGACCAAACC AGCTCCGCATCAGCAACTAC 130 ID1 NM_002165.3 GGTGGTGCGCTGTCTGTCT AGCACGTTTACCTGCTGCTC 104 ITGA2 NM_002203.3 TCAGGCTCACCGAGGTGACCAG GGTGCTGACCCAAAATGCCCTCTT 184 KRT7 NM_005556.3 ACACATCTGTGGTGCTGTCC CTCATACTGCGCCTTGACCT 150 MMP12 NM_002426.4 TGGTTTTTGCCCGTGGAGCTCAT GAATGGCCAATCTCGTGAACAGCA 186 MYC NM_002467.4 GGAGGCTATTCTGCCCATTT GGCTGCTGGTTTTCCACTAC 121 NANOS1 NM_199461.2 GAACAACAAGGAGGCGATG CGGGCAGTACTTGATGGTGT 144 OVOL1 NM_004561.3 CCGTGCGTCTCCACGTGCAA GGCTGTGGTGGGCAGAAGCC 100 OVOL2 NM_021220.3 CCGATGGACACCTGGCGACC GACGGTTCAGCATGCGCTGC 140 POU5F1 NM_002701.5 ATTCAGCCAAACGACCATCT AGCTTCCTCCACCCACTTCT 99 PRR9 NM_001195571.1 CCAAACAACCAGAAGGGAAA ACAGACCAGCCTGCATCAC 148 RP11-145M9.4 HG496359.1 GAACAGGCCCTTTTGCTGT TTTGGCAGCTATGCTGTTTG 149 SERPINE1 NM_000602.4 CTCTCTCTGCCCTCACCAAC CCAGGTTCTCTAGGGGCTTC 143 STAT1 NM_007315.3 GACCGCACCTTCAGTCTTTT TGAACTGGACCCCTGTCTTC 117 TFF1 NM_003225.2 CACCATGGAGAACAAGGTGA TGACACCAGGAAAACCACAA 133 TP63 NM_003722.4 GAAACGTACAGGCAACAGCA GATAAGCTGGCTCACAGAAGG 149 WIPF1 NM_003387.4 GACGGTCACCAATGACAGAA CTGGAACAATCCTCCCAGAC 171 ZNF702P NR_003578.1 CATGTGCTTGGCCATGTTAG ACCCTGGCTCCTTTTCTTTG 104 Primers used for conventional RT-PCR hOVOL1 NM_004561.3 CGCGGCGAGATCTACGTGCC GCCGCACGCCAGTGTGAGTT 438 mOvol1 NM_019935.3 TCGACTTACCAGAGCGAACC GTCTCGAAGGCTCATGTCCAA 244 mOvol2 NM_026924.3 CCACCCTGAGTGAAGAGGAG TTGCTGTCCCAAAAGAAGAGA 163 rOvol1 NM_001107572.1 CAGAGACCAGGGCTTCCTAC TGGGTTCGGACATGTCTCTTG 218 rOvol2 NM_001106519.1 CCTACAAATGCGAGGTGTGTT TGGGGATGTCAGCTTGTTCT 262

Renaud et al. www.pnas.org/cgi/content/short/1507397112 8of9 Table S4. Primers used for ChIP

T Gene name C /CGTTA Forward primer Reverse primer

ASCL2 −955 TCCACAAAAATCTCCGTTCC CACTGCAGAAATGCGAAAAA ERVFRD-1 −9,879 TGGAAGAGAAGACAAGTGCAAA ACACACACACCTGCATACACAC ERVFRD-1 −9,643 GGGGAATGAAGGAAGTAAAACC CACTTTGGGTCTGGGAATTGT ERVFRD-1 −8,976 TGGAAGACAGCCAGCTACATAA TGTCACCATGAAAATCTGAGAAA ERVFRD-1 −8,539 AGCCACCTGGTCTATGGTATTT GTTATACTGCACCTGGGCTTTT ERVFRD-1 −7,144 ATGTGCTTCAGTTTCCTGTTCT TTTTAGGCATTCACTTTAGCTTG ERVFRD-1 −7,040 CCATTCTCAAAGGGTATGTGGT CTTAGAGGGGAGGAGGTTGG ERVFRD-1 −3,193 CGAAATGCAACAAAATCCAC GACCTCAAGTGATCCACCTACC ERVFRD-1 −1,061 AGGAGGCAAAGGTTGCAGTGAG ATGAGAGTGGTTTCCAGCTTTTG ERVFRD-1 −663 AAATTAGCTGGGCATGGTGGTG CAATCTTGGCTCACTGCAACCT ERVFRD-1 −486 CAGAGCGAGGCTGTCTCCAAAA GCTCATGTCTGCCTGACTACCTAC ERVFRD-1 758 TGACTTACCATATGCTTTTCATCC CCTTAAGGGCTACAGAGTGAGGT ERVFRD-1 3,779 ACTGTTCACAGACCTTCCCATT AGCTAGAAGGATTCTTGGGCTGA ERVFRD-1 3,952 AAGAGGGATTTCAGCTTCTGACT GGTCCTGGACTAGCCTGTATTTT ERVFRD-1 4,318 CAGGACCTAAGTCTTGAGAGGAG GGACTAGATGTTGGATTCCCAGT ERVFRD-1 4,457 CGTGTCTGAAAACAGACTCCTAAA GGAAACTGAGTTAAGGGTGGAAC ERVFRD-1 4,853 ACACGTTTCTGGTTCTTCACACT ACTGTGCACTATTGTCACCTCCT ERVFRD-1 6,488 GTTCTCATTCTCACGCCTTCACT AGTAAGGGGATCCTGTACTTTGG ERVFRD-1 8,975 CTCCTGCTATAAAACTTGCCTTG ACACTGAGTCGGGTGTTAGTTTG ERVFRD-1 11,999 GCTTCTCTACCTCTACCCTCACC CCAAGCTCCTACAAATGAAACAC ERVFRD-1 13,414 GGACTAAGAAGCGACCACCA GGTGTTTTCTACCCCCACTATTC GAPDH – ACCCCCTTCCTTACAAGTGTTC ACAGGGGATCCATTCTTCTGGT ID1 −899 CGTCCGGGTTTTATGAATGG CCTGCAACAGTTCGTGATTTTT MYC −1,171 GGGAAAGAGGACCTGGAAAGG AGAGACAAATCCCCTTTGCGC ΔNP63 −1,812 AATGACTTTGGTAGGCAGTTGTG GCTAGGCGGTTCTGGATGAG

Renaud et al. www.pnas.org/cgi/content/short/1507397112 9of9