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Exon 2 Exon 3 STOP K8β intron Figure S1 …acacaagacagcugcagcag GUAUAGACGGGAAACAGGUGUCUAUCUUGGCCGGCUGGUUACUCAAAUGGGAACAAUGGCGCCACCUUGCUGUCUUUGUAG gcauuagaagaaaaggaugc…

Published byClaud Skinner Modified over 8 years ago

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Presentation on theme: "Exon 2 Exon 3 STOP K8β intron Figure S1 …acacaagacagcugcagcag GUAUAGACGGGAAACAGGUGUCUAUCUUGGCCGGCUGGUUACUCAAAUGGGAACAAUGGCGCCACCUUGCUGUCUUUGUAG gcauuagaagaaaaggaugc…"— Presentation transcript:

1 Exon 2 Exon 3 STOP K8β intron Figure S1 …acacaagacagcugcagcag GUAUAGACGGGAAACAGGUGUCUAUCUUGGCCGGCUGGUUACUCAAAUGGGAACAAUGGCGCCACCUUGCUGUCUUUGUAG gcauuagaagaaaaggaugc… 5’ss 3’ss oVM241 oVM242 oVM243 oVM244 oVM245 oVM246

2 K8β 1234 K8  - -++ORF57 RBM15 ++++ +--+ CNCNRNA CNCN GAPDH U6 -- -- K8 Figure S2

3 Figure S3 A B RBM15 SRSF3 ORF57 0100500 1000 β-tubulin ORF57 DNA (ng) HEK293 - full - cleaved VA (h) 063256 BCBL-1 1224 RBM15 SRSF3 β-tubulin ORF57 - full - cleaved SRSF3/RBM15 (%)418784722202478

4 Sample 1 Peptide Mass Peptide SequenceSequence Header 1512.67 1620.74 2861.37 1696.83 2286.16 EVSFQSTGESEWK MSGGWELELNGTEAK ITVTFNINNSIPPTFDGEEEPSQG QK AFVDFLSDEIKEER VEEQEPELTSTPNFVVEVIK >gi|4502491|ref|NP_001203.1| complement component 1, q subcomponent binding protein precursor [Homo sapiens]gi|730772|sp|Q07021.1|C1QBP_HUMAN RecName: Full=Complement component 1 Q subcomponent-binding protein, mitochondrial; AltName: Full=Glycoprot 1245.67 1453.68 1298.64 1141.64 IYVGNLPTDVR GSPHYFSPFRPY EKDLEDLFYK HGLVPFAFVR >gi|4506903|ref|NP_003760.1| splicing factor, arginine/serine-rich 9 [Homo sapiens]gi|114647286|ref|XP_001160785.1| PREDICTED: splicing factor, arginine/serine-rich 9 isoform 2 [Pan troglodytes]gi|3929377|sp|Q13242.1|SFRS9_HUMAN RecName: Full=Splic 1313.67 1505.77 1717.90 1618.79 LTPEEEEILNK ISSLLEEQFQQGK IIDVVYNASNNELVR QWYESHYALPLGR >gi|4506743|ref|NP_001003.1| ribosomal protein S8 [Homo sapiens]gi|6677813|ref|NP_033124.1| ribosomal protein S8 [Mus musculus]gi|13928986|ref|NP_113894.1| ribosomal protein S8 [Rattus norvegicus]gi|70778956|ref|NP_001020488.1| ribosomal protein S 1327.70 1031.58 1398.83 1270.74 ELLTLDEKDPR LFEGNALLR KQVVNIPSFIVR QVVNIPSFIVR >gi|14141193|ref|NP_001004.2| ribosomal protein S9 [Homo sapiens]gi|33504483|ref|NP_084043.1| ribosomal protein S9-like [Mus musculus]gi|110347598|ref|NP_112370.2| ribosomal protein S9 [Rattus norvegicus]gi|155372029|ref|NP_001094622.1| ribosomal 1129.61 1042.52 1620.76 2319.09 VRVELSNGEK AFGYYGPLR NPPGFAFVEFEDPR NPPGFAFVEFEDPRDAADAVR >gi|4506901|ref|NP_003008.1| splicing factor, arginine/serine-rich 3 [Homo sapiens]gi|8567402|ref|NP_038691.1| splicing factor, arginine/serine-rich 3 (SRp20) [Mus musculus]gi|77736345|ref|NP_001029872.1| splicing factor, arginine/serine-rich 3 [Bo 2077.02 1336.85 2366.27 AQQNNVEHKVETFSGVYK AIIIFVPVPQLK TLTAVHDAILEDLVFPSEIVGK >gi|4506741|ref|NP_001002.1| ribosomal protein S7 [Homo sapiens]gi|6755376|ref|NP_035430.1| ribosomal protein S7 [Mus musculus]gi|57618938|ref|NP_001009832.1| ribosomal protein S7 [Felis catus]gi|149642623|ref|NP_001092344.1| ribosomal protein S7 1302.61 2226.00 FSGSGSGTDFTLK DSTYSMSSTLTLTKDEYER >gi|1127148|pdb|1FLR|L Chain L, 4-4-20 Fab Fragment 1941.92 VWLDPNETNEIANANSR >gi|4506609|ref|NP_000972.1| ribosomal protein L19 [Homo sapiens]gi|14389297|ref|NP_112365.1| ribosomal protein L19 [Rattus norvegicus]gi|94966831|ref|NP_001035606.1| ribosomal protein L19 [Bos taurus]gi|197100875|ref|NP_001125553.1| ribosomal pro 1344.72 TAVVVGTITDDVR >gi|4506607|ref|NP_000970.1| ribosomal protein L18 [Homo sapiens]gi|109125415|ref|XP_001114009.1| PREDICTED: similar to ribosomal protein L18 [Macaca mulatta]gi|548749|sp|Q07020.2|RL18_HUMAN RecName: Full=60S ribosomal protein L18gi|75076040|sp|Q4 1777.96 GLDVDSLVIEHIQVNK >gi|42542645|gb|AAH66324.1| Ribosomal protein L17 [Homo sapiens] 1545.84 VLEQLTGQTPVFSK >gi|495126|emb|CAA55816.1| ribosomal protein L11 [Homo sapiens] Sample2 1512.67 1620.74 1696.83 2286.16 EVSFQSTGESEWK MSGGWELELNGTEAK AFVDFLSDEIKEER VEEQEPELTSTPNFVVEVIK >gi|4502491|ref|NP_001203.1| complement component 1, q subcomponent binding protein precursor [Homo sapiens]gi|730772|sp|Q07021.1|C1QBP_HUMAN RecName: Full=Complement component 1 Q subcomponent-binding protein, mitochondrial; AltName: Full=Glycoprot 1313.67 1505.77 1717.90 1618.79 LTPEEEEILNK ISSLLEEQFQQGK IIDVVYNASNNELVR QWYESHYALPLGR >gi|4506743|ref|NP_001003.1| ribosomal protein S8 [Homo sapiens]gi|6677813|ref|NP_033124.1| ribosomal protein S8 [Mus musculus]gi|13928986|ref|NP_113894.1| ribosomal protein S8 [Rattus norvegicus]gi|70778956|ref|NP_001020488.1| ribosomal protein S 1302.61 1590.73 2226.00 FSGSGSGTDFTLK QNGVLNSWTDQDSK DSTYSMSSTLTLTKDEYER >gi|1127148|pdb|1FLR|L Chain L, 4-4-20 Fab Fragment 1652.74 1245.67 1453.68 FEDPRDAEDAIYGR IYVGNLPTDVR GSPHYFSPFRPY >gi|4506903|ref|NP_003760.1| splicing factor, arginine/serine-rich 9 [Homo sapiens]gi|114647286|ref|XP_001160785.1| PREDICTED: splicing factor, arginine/serine-rich 9 isoform 2 [Pan troglodytes]gi|3929377|sp|Q13242.1|SFRS9_HUMAN RecName: Full=Splic 1941.92 VWLDPNETNEIANANSR >gi|4506609|ref|NP_000972.1| ribosomal protein L19 [Homo sapiens]gi|14389297|ref|NP_112365.1| ribosomal protein L19 [Rattus norvegicus]gi|94966831|ref|NP_001035606.1| ribosomal protein L19 [Bos taurus]gi|197100875|ref|NP_001125553.1| ribosomal pro 1270.74 QVVNIPSFIVR >gi|14141193|ref|NP_001004.2| ribosomal protein S9 [Homo sapiens]gi|33504483|ref|NP_084043.1| ribosomal protein S9-like [Mus musculus]gi|110347598|ref|NP_112370.2| ribosomal protein S9 [Rattus norvegicus]gi|155372029|ref|NP_001094622.1| ribosomal 1344.72 TAVVVGTITDDVR >gi|4506607|ref|NP_000970.1| ribosomal protein L18 [Homo sapiens]gi|109125415|ref|XP_001114009.1| PREDICTED: similar to ribosomal protein L18 [Macaca mulatta]gi|548749|sp|Q07020.2|RL18_HUMAN RecName: Full=60S ribosomal protein L18gi|75076040|sp|Q4 1777.96 GLDVDSLVIEHIQVNK >gi|42542645|gb|AAH66324.1| Ribosomal protein L17 [Homo sapiens] 2319.09 NPPGFAFVEFEDPRDAADAVR >gi|4506901|ref|NP_003008.1| splicing factor, arginine/serine-rich 3 [Homo sapiens]gi|8567402|ref|NP_038691.1| splicing factor, arginine/serine-rich 3 (SRp20) [Mus musculus]gi|77736345|ref|NP_001029872.1| splicing factor, arginine/serine-rich 3 [Bos taurus] Supplementary Table S1: Peptides and corresponding proteins associated with KSHV ORF57 in THE absence (Fig 2A, sample 1) or presence (Fig 2A, sample 2) of ectopically expressed RBM15. The peptides and proteins were identified with LC-MS/MS.

5 OligoSequencePosition KSHV oVM241 5'-Biotin-rArCrArCrArArGrArCrArGrCrUrGrCrArGrCrArGrGrUrArUrAdTdT-3'75544-75568 oVM242 5'-Biotin-rGrUrArUrArGrArCrGrGrGrArArArCrArGrGrUrGrUrCrUrArUdTdT-3'75564-75588 oVM243 5'-Biotin-rUrCrUrArUrCrUrUrGrGrCrCrGrGrCrUrGrGrUrUrArCrUrCrAdTdT-3'75584-75608 oVM244 5'-Biotin-rArCrUrCrArArArUrGrGrGrArArCrArArUrGrGrCrGrCrCrArCdTdT-3'75604-75628 oVM245 5'-Biotin-rGrCrCrArCrCrUrUrGrCrUrGrUrCrUrUrUrGrUrArGrGrCrArUdTdT-3'75624-75648 oVM246 5'-Biotin-rUrGrUrArGrGrCrArUrUrArGrArArGrArArArArGrGrArUrGrCdTdT-3'75640-75664 HPV16 oJR9 (wt) 5′-Biotin-rCrCrArGrArCrArCrCrGrGrArArArCrCrCrCrUrGrCrCrArCrArCrCrArC-3′3488-3516 oJR10 (mt) 5′-Biotin-rCrCrArGrArUrGrUrCrGrGrArArArCrCrUrCrUrGrCrUrGrUrGrCrUrGrU-3′3488-3516 SUPPLEMENTARY TABLE S2. Biotinylated RNA oligos from the KSHV K8β intron region and the HPV16 ESE used in RNA pulldowns. Underlined are point mutations.

6 Supplementary Table S3: Primers used for RT-PCR and Northern Blotting. T7, T7 promoter; U1bs, U1 binding site OligoSequenceTargetPosition oST15'-TAATACGACTCACTATAGGG /ACCACCAAGAGGACCACACATTTC-3' T7/KSHV K8 (exon 1) 75182-75205 oST35'-GTACTCACCCC/CACACAAA GTCTGGCATGGTTCTCCC-3' U1bs/ KSHV K8 (exon 3) 75838-75813 oVM2135'-CCCGGGATCCACCGGTCGCCACCA-3'BamHI/GFP656-679 oVM1625'-GGTTCAGGGGGAGGTGTGG-3'GFP1486-1468 oZMZ2465’-CAATGGGACGCGTGCAAAGC-3’BPV-1 (Pr1)7345-7364 oZMZ2495’-TAATACGACTCACTATAGGGA/ CAGTATTTGTGCTTGTCCTT-3’ T7/BPV-1 (Pr2)3738-3719 oXFL0495’-TAATACGACTCACTATAGG /TTTAATGCTTTGCTTGATGC-3’ T7/BPV-1 (Pr3)5963-5982 oXFL0465’-CATATCCCCATCTTCTATGTGC-3’BPV-1 (Pr4)6196-6175 oSB235’-TATTAGGCAGCACTTGGC-3’HPV16 (Pr1)3385-3402 oXHW415’-CAACATATTCATCCGTGCTTACAAC-3’HPV16 (Pr2)5723-5699 oST2855’-TGGCTTTGGTGCTATGGACT-3’HPV16 (Pr3)6247-6266 oYL95’-GAAGTAGATATGGCAGCACA-3’HPV16 (Pr4)6690-6671 oZMZ2695'-GTCATCAATGGAAATCCCATCACC-3'GAPDH202-225 oZMZ2705'-TGAGTCCTTCCACGATACCAAA-3'GAPDH523-502 oST32CGGACGACGGCACAACCAKSHV K8 (exon 3)75813-75796 oST197AAAATATGGAACGCTTCACGAU6106-86

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