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oVM246. 3’ss. oVM245. Exon 3. oVM244. K8 β intron. …acacaagacagcugcagcag GUAUAGACGGGAAACAGGUGUCUAUCUUGGCCGGCUGGUUACUCAAAUGGGAACAAUGGCGCCACCUUGCUGUCUUUGUAG gcauuagaagaaaaggaugc…. oVM243. Exon 2. oVM242. STOP. 5’ss. oVM241. Figure S1. K8 β. 1. 2. 3. 4. RBM15. -. -. +. +. -. - PowerPoint PPT Presentation
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Ex
on
2E
xo
n 3
ST
OP
K8
β i
ntr
on
Figure S1
…ac
acaa
gaca
gcug
cagc
agG
UA
UA
GA
CG
GG
AA
AC
AG
GU
GU
CU
AU
CU
UG
GC
CG
GC
UG
GU
UA
CU
CA
AA
UG
GG
AA
CA
AU
GG
CG
CC
AC
CU
UG
CU
GU
CU
UU
GU
AG
gcau
uaga
agaa
aagg
augc
…
5’ss
3’ss
oV
M2
41o
VM
242
oV
M2
43o
VM
244
oV
M2
45o
VM
246
K8β 1 2 3 4
K8-- + +ORF57
RBM15
++ + +
+- - +
C N C NRNA C N C N
GAPDH
U6
--K8
Figure S2
Figure S3
A
B
RBM15
SRSF3
ORF57
0 100
500
1000
β-tubulin
ORF57 DNA (ng)
HEK293
- full- cleaved
VA (h) 0 6 32 56
BCBL-1
12 24
RBM15
SRSF3
β-tubulin
ORF57- full- cleaved
SRSF3/RBM15 (%) 41 87847 2220 2478
Sample 1Peptide Mass
Peptide Sequence Sequence Header
1512.671620.742861.371696.832286.16
EVSFQSTGESEWKMSGGWELELNGTEAKITVTFNINNSIPPTFDGEEEPSQGQKAFVDFLSDEIKEER 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
1245.671453.681298.641141.64
IYVGNLPTDVR GSPHYFSPFRPYEKDLEDLFYKHGLVPFAFVR
>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
1313.671505.771717.901618.79
LTPEEEEILNKISSLLEEQFQQGKIIDVVYNASNNELVRQWYESHYALPLGR
>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
1327.701031.581398.831270.74
ELLTLDEKDPRLFEGNALLRKQVVNIPSFIVRQVVNIPSFIVR
>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
1129.611042.521620.762319.09
VRVELSNGEKAFGYYGPLRNPPGFAFVEFEDPRNPPGFAFVEFEDPRDAADAVR
>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
2077.021336.852366.27
AQQNNVEHKVETFSGVYKAIIIFVPVPQLKTLTAVHDAILEDLVFPSEIVGK
>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
1302.612226.00
FSGSGSGTDFTLKDSTYSMSSTLTLTKDEYER
>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
1512.671620.741696.832286.16
EVSFQSTGESEWKMSGGWELELNGTEAKAFVDFLSDEIKEERVEEQEPELTSTPNFVVEVIK
>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
1313.671505.771717.901618.79
LTPEEEEILNKISSLLEEQFQQGKIIDVVYNASNNELVRQWYESHYALPLGR
>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
1302.611590.731590.732226.00
FSGSGSGTDFTLKQNGVLNSWTDQDSKQNGVLNSWTDQDSKDSTYSMSSTLTLTKDEYER
>gi|1127148|pdb|1FLR|L Chain L, 4-4-20 Fab Fragment
1652.741245.671453.68
FEDPRDAEDAIYGRIYVGNLPTDVRGSPHYFSPFRPY
>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.
Oligo Sequence Position
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' 75624-75648oVM246 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 andthe HPV16 ESE used in RNA pulldowns. Underlined are point mutations.
Supplementary Table S3: Primers used for RT-PCR and Northern Blotting.
T7, T7 promoter; U1bs, U1 binding site
Oligo Sequence Target Position
oST1 5'-TAATACGACTCACTATAGGG/ACCACCAAGAGGACCACACATTTC-3'
T7/KSHV K8 (exon 1)
75182-75205
oST3 5'-GTACTCACCCC/CACACAAAGTCTGGCATGGTTCTCCC-3'
U1bs/ KSHV K8 (exon 3)
75838-75813
oVM213 5'-CCCGGGATCCACCGGTCGCCACCA-3' BamHI/GFP 656-679
oVM162 5'-GGTTCAGGGGGAGGTGTGG-3' GFP 1486-1468
oZMZ246 5’-CAATGGGACGCGTGCAAAGC-3’ BPV-1 (Pr1) 7345-7364
oZMZ249 5’-TAATACGACTCACTATAGGGA/CAGTATTTGTGCTTGTCCTT-3’
T7/BPV-1 (Pr2) 3738-3719
oXFL049 5’-TAATACGACTCACTATAGG/TTTAATGCTTTGCTTGATGC-3’
T7/BPV-1 (Pr3) 5963-5982
oXFL046 5’-CATATCCCCATCTTCTATGTGC-3’ BPV-1 (Pr4) 6196-6175
oSB23 5’-TATTAGGCAGCACTTGGC-3’ HPV16 (Pr1) 3385-3402
oXHW41 5’-CAACATATTCATCCGTGCTTACAAC-3’ HPV16 (Pr2) 5723-5699
oST285 5’-TGGCTTTGGTGCTATGGACT-3’ HPV16 (Pr3) 6247-6266
oYL9 5’-GAAGTAGATATGGCAGCACA-3’ HPV16 (Pr4) 6690-6671
oZMZ269 5'-GTCATCAATGGAAATCCCATCACC-3' GAPDH 202-225
oZMZ270 5'-TGAGTCCTTCCACGATACCAAA-3' GAPDH 523-502
oST32 CGGACGACGGCACAACCA KSHV K8 (exon 3) 75813-75796
oST197 AAAATATGGAACGCTTCACGA U6 106-86
