>EIF4H|NM_022170|3'UTR
   1 GCCTGCGGTTGGGAGGGAATGGGGCGTGGGGGGTTAGAGCAGGACCACAGCCTGGTGAGTCCCCGGGCAGCCGTCCTGCA
  81 GCCGCCACTCCTGCGCCTGCCATTGGCCTCCTCACAGCGGAAACACAGCTTGTGAGTGCATGTCAGCTGTTAACAAGTGG
 161 TTTTTAGTACATTCTGGGCTTTGCTGTATCTATCTAGTGCCTGTTTGTGCGTTTTTTTCTTTCTTCCGCTGCTTCCCCAT
 241 TTTCCTTCTGTCCTTTTTCTCCTGCTCCTTGTTTTCCCAGCAGCACATGGGGTTCCTCGGAGGAGCAGAGGTGGCCGCCG
 321 TGGGGGGGCGTTTGGGCTGCGGTGCTGCGTCATTTTTCCTTTGCTTTCTCTTTACTTTAGACACTGGCCCAACTCCAGGC
 401 GTTTCCTTTCATTCCCTCAGTGCTTCTCTTCTGACCTGCATGTTGAGTTCTGTATTGCTGGGGCTTCCAACAAAAACCAG
 481 AGTCACTGACAGAGGGAACAGCAGAGACCTTGTTGGTATTCAGCTGTGATGGATATAGAGAATCAGAGGCACCTTGTTTT
 561 CACAACTAGGATAAAAATATCTGCAGGGTCCTTTCCATTCCTATTTAGAGGGAGTCCTGGCTCCATGACCCCCTCCCGAG
 641 TGGACTGTCCAAGCAGATAGGCTCACACGAGAAACAGTGAGGCTGAAAGGGGGGGCTATGGAAGAGCGGTAGGGAGTCCA
 721 CGGAGAAGATGCAGTGAATGCTTGCATGCATTCACACGTGTGTGTGTCCCAGCTAGTTCACTCCTTTCGCCGTGCGTGGT
 801 GGAGGCTGGCCTCTCTGGCTGGGTGCAGTGAATGGCCAGCGGGTTTCTTTTCTGCTGGGCCAAGGCGCTTTGGGGGTGGA
 881 GGGGGTGGTGCTGGTGCTGCACTGGGCTGACTGCGGCGCTGACGCAGCGTTTCCCCCCATCCCTGTTGCCTGTGTGTTGT
 961 GTGGATCTGTTCCTAGTATAGGCAACATAATGAGATACTGTGCTTCCCACCTCCCCTTCAGTTCAGAGCCAAAATGGGTC
1041 TAGAATCTGGCACTTTACTCATTTCCTTTGATAAATTGTACTATGCAGAGCTGTCAGGAACCTTCAGATAGCAGTAGAGG
1121 ACTGCAGCTGTCTAGGTCTGCGGCCACATCTTGGGGACACACTGGACTGTTCCCATGTGCAGGGTTCAGCAGTTATGTGG
1201 GAGTGCTAGGGGTTAGGCTTTTGAGCTTGAACGCCTGCGTGTGAACAGATGAAAAATCCTTCAGTACCCAAGTCCCAGTC
1281 TGTCCTATGGGGAGCAGTTTGGGGGCGGCCGGCAGCAGGAGCCTGGGAAAGAGGCCCTCGCCAGGTGATGGCAGGGCCAG
1361 GGTGGCCTGGGGCACCCAGCGGAATGTGCTTAGTATTTGGTCACCAGCCGTCATCCTGGGCTTTTCCTACTGTGTCTTGT
1441 TACAAGGCCTCAGCAATCCACAGAACTCTCTCTCCTTCCTTCCACCTGTCAGCTTCTCTGCTTCTGAGATAAGAACCATT
1521 TGTGTAACACCAACACTTAACTTCAGAAAGACATGCATTATGTGGTGTAATCAAACCCGATGCTTTCAGATGACCTACTT
1601 ACATCTTCAATGTGGATAAGATAAAGAACAAAACACATGCATCTAAACTGCTGGGCAATCCAGTTGACTTTTAAATGTAA
1681 GAATGGAATTCCAAACACTTAACACATTCAGCTATATGACAGAAAGTAAATCTATGGATATGGTATTTTGTGAATGATCT
1761 TTTAAATAAAAGAAAACCTTACGTAATATTT

ID	Duplex structure	Position	Score	MFE
1	miRNA 3' guGAGUCGCCGGCAAAGGGGUu 5' :|| | ||:| ||||||| Target 5' ctTTCTTCCGCTGCTTCCCCAt 3'	219 - 240	160.00	-17.20
2	miRNA 3' guGAGU-CGCCGGCAAAGGGGuu 5' || | || | ||||||||| Target 5' cgCTGACGCAG-CGTTTCCCCcc 3'	917 - 938	143.00	-19.80
3	miRNA 3' guGA-GUCGCCG-GCAAAGGGGUu 5' || | || | :||||:|||| Target 5' ttCTCCTGCTCCTTGTTTTCCCAg 3'	257 - 280	140.00	-17.50

Mutant ID	Mutant Position	Mutant Source
COSN30506350	8	COSMIC
COSN14300882	26	COSMIC
COSN30128096	30	COSMIC
COSN30495418	65	COSMIC
COSN31531541	66	COSMIC
COSN30499239	141	COSMIC
COSN31523115	170	COSMIC
COSN23147096	286	COSMIC
COSN32073891	321	COSMIC
COSN30606725	329	COSMIC
COSN2234626	368	COSMIC
COSN19657016	401	COSMIC
COSN30703376	401	COSMIC
COSN26585833	458	COSMIC
COSN20344171	695	COSMIC
COSN15763972	817	COSMIC
COSN30545151	953	COSMIC
COSN31546031	976	COSMIC
COSN31486972	1048	COSMIC
COSN30545207	1086	COSMIC
COSN30158034	1089	COSMIC
COSN31549529	1748	COSMIC
COSN26279660	1780	COSMIC

Mutant ID	Mutant Position	Mutant Source
rs1447066597	4	dbSNP
rs1375628741	7	dbSNP
rs782454237	8	dbSNP
rs782576980	19	dbSNP
rs782302162	25	dbSNP
rs782192917	26	dbSNP
rs782406670	26	dbSNP
rs535742148	27	dbSNP
rs782449959	28	dbSNP
rs782220493	29	dbSNP
rs557645944	30	dbSNP
rs781929810	31	dbSNP
rs782045444	32	dbSNP
rs782422377	33	dbSNP
rs1403484370	35	dbSNP
rs1411755730	41	dbSNP
rs575700661	42	dbSNP
rs782127280	47	dbSNP
rs1180509928	48	dbSNP
rs1440564472	50	dbSNP
rs546032982	51	dbSNP
rs782667557	53	dbSNP
rs563827396	56	dbSNP
rs1489700851	65	dbSNP
rs1266354891	66	dbSNP
rs1320374895	67	dbSNP
rs1287564941	69	dbSNP
rs572635003	73	dbSNP
rs555278621	74	dbSNP
rs575358681	84	dbSNP
rs1368064384	85	dbSNP
rs1434212880	87	dbSNP
rs1346835243	90	dbSNP
rs1159166914	91	dbSNP
rs1420454180	94	dbSNP
rs1379734297	95	dbSNP
rs1157106019	96	dbSNP
rs1435089329	97	dbSNP
rs1268499263	98	dbSNP
rs1190917956	103	dbSNP
rs1487704403	104	dbSNP
rs1266061180	108	dbSNP
rs1217955211	109	dbSNP
rs1352406823	110	dbSNP
rs561497484	114	dbSNP
rs1234995896	117	dbSNP
rs1312340390	119	dbSNP
rs1300343611	123	dbSNP
rs1217321215	133	dbSNP
rs1342135094	140	dbSNP
rs1298774617	165	dbSNP
rs1399272300	172	dbSNP
rs1360020008	178	dbSNP
rs1287122881	187	dbSNP
rs1467788433	187	dbSNP
rs1378489390	191	dbSNP
rs1175626703	195	dbSNP
rs1426206354	202	dbSNP
rs1431844709	202	dbSNP
rs1195058778	206	dbSNP
rs1488225944	207	dbSNP
rs1260409416	210	dbSNP
rs1201612826	211	dbSNP
rs1255121889	212	dbSNP
rs1483283060	212	dbSNP
rs1230571985	221	dbSNP
rs528897220	228	dbSNP
rs1278564798	229	dbSNP
rs1217942859	233	dbSNP
rs1338260999	236	dbSNP
rs1314755497	237	dbSNP
rs1397932666	241	dbSNP
rs782705400	249	dbSNP
rs1302539923	257	dbSNP
rs1468389231	268	dbSNP
rs1171750800	270	dbSNP
rs1477786470	275	dbSNP
rs1422175355	276	dbSNP
rs1190215731	278	dbSNP
rs544136216	287	dbSNP
rs562848775	289	dbSNP
rs187036467	297	dbSNP
rs1458769107	299	dbSNP
rs1258481490	300	dbSNP
rs1197843498	306	dbSNP
rs1315836673	318	dbSNP
rs1267194745	319	dbSNP
rs1245402751	320	dbSNP
rs1312041115	321	dbSNP
rs1332276468	322	dbSNP
rs1379316417	322	dbSNP
rs1465493490	323	dbSNP
rs551636758	328	dbSNP
rs1367388245	329	dbSNP
rs1181152620	331	dbSNP
rs1440985551	334	dbSNP
rs1177595853	335	dbSNP
rs1456316503	336	dbSNP
rs537769003	337	dbSNP
rs1210394774	341	dbSNP
rs1355469633	342	dbSNP
rs1264949418	343	dbSNP
rs1243941495	349	dbSNP
rs1355939584	350	dbSNP
rs528571636	355	dbSNP
rs782521136	363	dbSNP
rs1446298528	371	dbSNP
rs1334373923	372	dbSNP
rs1325867254	373	dbSNP
rs547088922	375	dbSNP
rs1390718981	380	dbSNP
rs1162218501	382	dbSNP
rs1421034843	385	dbSNP
rs1380357944	390	dbSNP
rs55815268	394	dbSNP
rs1157704699	397	dbSNP
rs1248243222	400	dbSNP
rs1199227719	401	dbSNP
rs1265260458	403	dbSNP
rs568324188	405	dbSNP
rs1489339486	413	dbSNP
rs535843720	416	dbSNP
rs1239733056	421	dbSNP
rs557364764	424	dbSNP
rs1276815713	425	dbSNP
rs1234510830	432	dbSNP
rs1330854557	436	dbSNP
rs1301892873	441	dbSNP
rs782071929	441	dbSNP
rs1402348685	442	dbSNP
rs1360647858	444	dbSNP
rs1296972524	471	dbSNP
rs1398604509	477	dbSNP
rs1426897041	479	dbSNP
rs764267904	480	dbSNP
rs1477595969	488	dbSNP
rs1426913260	497	dbSNP
rs1195782458	517	dbSNP
rs569247953	520	dbSNP
rs1264003266	525	dbSNP
rs1445351033	527	dbSNP
rs774618863	534	dbSNP
rs1201229617	537	dbSNP
rs1344782144	538	dbSNP
rs1281733905	547	dbSNP
rs1221013395	548	dbSNP
rs1341611642	554	dbSNP
rs1428121081	559	dbSNP
rs1360355849	567	dbSNP
rs1334457500	578	dbSNP
rs1413503768	581	dbSNP
rs539625737	591	dbSNP
rs1172467941	602	dbSNP
rs1478019230	607	dbSNP
rs1184678043	617	dbSNP
rs1422913057	617	dbSNP
rs1474496851	621	dbSNP
rs1239237297	630	dbSNP
rs1188550062	633	dbSNP
rs557958107	634	dbSNP
rs1200976957	639	dbSNP
rs1319024880	648	dbSNP
rs1272971376	656	dbSNP
rs1213298787	663	dbSNP
rs1380737236	670	dbSNP
rs1284230954	675	dbSNP
rs1446841615	676	dbSNP
rs1380045636	678	dbSNP
rs1332946970	685	dbSNP
rs1491474929	688	dbSNP
rs1491427393	689	dbSNP
rs558005791	689	dbSNP
rs1406980624	690	dbSNP
rs1163858057	691	dbSNP
rs1180707358	692	dbSNP
rs573123037	693	dbSNP
rs1232614653	695	dbSNP
rs1203070817	697	dbSNP
rs35927562	698	dbSNP
rs539906540	708	dbSNP
rs1289985861	709	dbSNP
rs1244691063	711	dbSNP
rs1356603370	719	dbSNP
rs1311064994	722	dbSNP
rs555448696	723	dbSNP
rs1375922460	731	dbSNP
rs1296173012	736	dbSNP
rs1438545297	740	dbSNP
rs1350461572	742	dbSNP
rs1163981236	758	dbSNP
rs1383434432	758	dbSNP
rs1424138701	758	dbSNP
rs1160703246	760	dbSNP
rs573554269	765	dbSNP
rs1407719699	766	dbSNP
rs1175789431	767	dbSNP
rs1489663261	781	dbSNP
rs1247845569	785	dbSNP
rs544236987	789	dbSNP
rs562625826	790	dbSNP
rs1240432239	792	dbSNP
rs533293491	793	dbSNP
rs545347649	797	dbSNP
rs560357870	800	dbSNP
rs1304402390	811	dbSNP
rs1220799516	823	dbSNP
rs1367998214	828	dbSNP
rs34460100	834	dbSNP
rs115874427	840	dbSNP
rs868951948	841	dbSNP
rs1300039684	842	dbSNP
rs1420552200	850	dbSNP
rs1344888939	856	dbSNP
rs546733200	862	dbSNP
rs1435331108	863	dbSNP
rs1426254489	867	dbSNP
rs1199371558	868	dbSNP
rs34839061	874	dbSNP
rs1264725723	875	dbSNP
rs1216781120	877	dbSNP
rs1258623600	878	dbSNP
rs1485259234	878	dbSNP
rs1211767533	880	dbSNP
rs568584758	882	dbSNP
rs529260836	883	dbSNP
rs35864042	886	dbSNP
rs1282351198	887	dbSNP
rs1221614239	893	dbSNP
rs1278749150	895	dbSNP
rs1342230679	895	dbSNP
rs35689317	896	dbSNP
rs1382913653	899	dbSNP
rs1384319696	907	dbSNP
rs1287358590	910	dbSNP
rs1467739103	915	dbSNP
rs868917281	916	dbSNP
rs1176092941	918	dbSNP
rs1426499963	922	dbSNP
rs1427455111	924	dbSNP
rs35239215	925	dbSNP
rs1183266290	926	dbSNP
rs1483333803	927	dbSNP
rs577983075	929	dbSNP
rs1201597524	930	dbSNP
rs1319730995	933	dbSNP
rs550985054	937	dbSNP
rs1226542671	942	dbSNP
rs1325962496	949	dbSNP
rs1293443018	950	dbSNP
rs1398022853	955	dbSNP
rs1383881046	964	dbSNP
rs1336322518	970	dbSNP
rs569183773	975	dbSNP
rs1450971954	986	dbSNP
rs1402877720	993	dbSNP
rs1169017622	995	dbSNP
rs1461882754	1000	dbSNP
rs373695868	1011	dbSNP
rs1166213207	1012	dbSNP
rs1423874203	1014	dbSNP
rs1242219931	1017	dbSNP
rs1181336264	1031	dbSNP
rs1458943722	1033	dbSNP
rs1238519713	1035	dbSNP
rs1195163454	1037	dbSNP
rs1337304080	1039	dbSNP
rs1267134129	1040	dbSNP
rs1247171847	1043	dbSNP
rs1360223104	1054	dbSNP
rs1314686415	1057	dbSNP
rs1379610861	1062	dbSNP
rs1450654308	1062	dbSNP
rs558044327	1065	dbSNP
rs1329880897	1077	dbSNP
rs34251399	1084	dbSNP
rs1395038602	1088	dbSNP
rs1295796989	1092	dbSNP
rs1424790469	1107	dbSNP
rs1384025790	1114	dbSNP
rs1161317353	1116	dbSNP
rs34582986	1132	dbSNP
rs1441087074	1142	dbSNP
rs1364179646	1143	dbSNP
rs1180896780	1152	dbSNP
rs1435865845	1157	dbSNP
rs149967248	1159	dbSNP
rs534407152	1162	dbSNP
rs35628518	1168	dbSNP
rs1210341406	1172	dbSNP
rs1490539560	1173	dbSNP
rs1291077336	1185	dbSNP
rs35529759	1189	dbSNP
rs1356049504	1191	dbSNP
rs35668946	1197	dbSNP
rs1238867343	1205	dbSNP
rs1334318476	1215	dbSNP
rs1304115018	1221	dbSNP
rs1406048467	1223	dbSNP
rs1364413245	1227	dbSNP
rs555162045	1234	dbSNP
rs11557834	1236	dbSNP
rs573715163	1239	dbSNP
rs1468450699	1240	dbSNP
rs1379108284	1270	dbSNP
rs1200105490	1278	dbSNP
rs1480033133	1279	dbSNP
rs1220431167	1288	dbSNP
rs1489310879	1295	dbSNP
rs1285269969	1306	dbSNP
rs1204750730	1307	dbSNP
rs1349024133	1308	dbSNP
rs1276906790	1310	dbSNP
rs1224674104	1311	dbSNP
rs537422603	1312	dbSNP
rs1281923101	1324	dbSNP
rs1401347639	1326	dbSNP
rs1326479463	1328	dbSNP
rs1317222094	1329	dbSNP
rs1398556967	1330	dbSNP
rs1404096013	1331	dbSNP
rs1176818985	1336	dbSNP
rs1469091333	1337	dbSNP
rs1427200721	1339	dbSNP
rs1195716042	1340	dbSNP
rs556167786	1341	dbSNP
rs1192752738	1346	dbSNP
rs1445437011	1349	dbSNP
rs1265216578	1361	dbSNP
rs577730616	1363	dbSNP
rs1323234445	1367	dbSNP
rs13244093	1371	dbSNP
rs1220943937	1374	dbSNP
rs1339524704	1381	dbSNP
rs879977642	1382	dbSNP
rs1341957722	1383	dbSNP
rs1336978544	1386	dbSNP
rs1451702501	1407	dbSNP
rs1403606448	1410	dbSNP
rs1172759440	1411	dbSNP
rs545285956	1414	dbSNP
rs1420331809	1432	dbSNP
rs1167466129	1440	dbSNP
rs1474463648	1441	dbSNP
rs1245241045	1444	dbSNP
rs1461160436	1449	dbSNP
rs1241637668	1464	dbSNP
rs1207256141	1466	dbSNP
rs1438844702	1466	dbSNP
rs1272911262	1468	dbSNP
rs1212361318	1469	dbSNP
rs1360939704	1470	dbSNP
rs1315369175	1471	dbSNP
rs1244727035	1473	dbSNP
rs1380347265	1478	dbSNP
rs1333050219	1484	dbSNP
rs1442644807	1491	dbSNP
rs1353930201	1499	dbSNP
rs1326732784	1525	dbSNP
rs1408325105	1531	dbSNP
rs781847009	1531	dbSNP
rs1164481996	1535	dbSNP
rs1444170634	1537	dbSNP
rs1383796324	1554	dbSNP
rs1179382369	1557	dbSNP
rs1481163567	1561	dbSNP
rs1203010628	1566	dbSNP
rs1490703773	1567	dbSNP
rs1221297209	1568	dbSNP
rs1356900721	1576	dbSNP
rs560466954	1579	dbSNP
rs1224167141	1583	dbSNP
rs11557832	1585	dbSNP
rs1372313148	1590	dbSNP
rs1277722183	1594	dbSNP
rs1438466905	1594	dbSNP
rs1367531812	1601	dbSNP
rs1303780084	1603	dbSNP
rs1424088272	1604	dbSNP
rs782690308	1606	dbSNP
rs78099872	1611	dbSNP
rs1161517493	1612	dbSNP
rs1420106578	1613	dbSNP
rs1407656852	1617	dbSNP
rs1176018145	1622	dbSNP
rs1481667491	1623	dbSNP
rs542846380	1627	dbSNP
rs1270215312	1628	dbSNP
rs1198146341	1630	dbSNP
rs1489619420	1632	dbSNP
rs1262174551	1633	dbSNP
rs183678176	1637	dbSNP
rs1316150382	1638	dbSNP
rs1281796276	1641	dbSNP
rs1346141932	1659	dbSNP
rs201082008	1663	dbSNP
rs1404457590	1678	dbSNP
rs1365551186	1684	dbSNP
rs1290879839	1694	dbSNP
rs1454467085	1694	dbSNP
rs1344979502	1696	dbSNP
rs1157450219	1700	dbSNP
rs1471158348	1702	dbSNP
rs1430660557	1716	dbSNP
rs1199301397	1717	dbSNP
rs762221385	1721	dbSNP
rs1479181292	1726	dbSNP
rs1263168970	1729	dbSNP
rs367939007	1735	dbSNP
rs1186743411	1739	dbSNP
rs1485305789	1741	dbSNP
rs1239994544	1742	dbSNP
rs1205406023	1746	dbSNP
rs1323955260	1755	dbSNP
rs1262489051	1756	dbSNP
rs1221560955	1768	dbSNP
rs190525159	1772	dbSNP
rs1297729474	1783	dbSNP
rs1361606405	1784	dbSNP
rs1318749569	1787	dbSNP

- mBio, 2013

UNLABELLED: The question of how HIV-1 interfaces with cellular microRNA (miRNA) biogenesis and effector mechanisms has been highly controversial. Here, we first used deep sequencing of small RNAs present in two different infected cell lines (TZM-bl and C8166) and two types of primary human cells (CD4(+) peripheral blood mononuclear cells [PBMCs] and macrophages) to unequivocally demonstrate that HIV-1 does not encode any viral miRNAs. Perhaps surprisingly, we also observed that infection of T cells by HIV-1 has only a modest effect on the expression of cellular miRNAs at early times after infection. Comprehensive analysis of miRNA binding to the HIV-1 genome using the photoactivatable ribonucleoside-induced cross-linking and immunoprecipitation (PAR-CLIP) technique revealed several binding sites for cellular miRNAs, a subset of which were shown to be capable of mediating miRNA-mediated repression of gene expression. However, the main finding from this analysis is that HIV-1 transcripts are largely refractory to miRNA binding, most probably due to extensive viral RNA secondary structure. Together, these data demonstrate that HIV-1 neither encodes viral miRNAs nor strongly influences cellular miRNA expression, at least early after infection, and imply that HIV-1 transcripts have evolved to avoid inhibition by preexisting cellular miRNAs by adopting extensive RNA secondary structures that occlude most potential miRNA binding sites. IMPORTANCE: MicroRNAs (miRNAs) are a ubiquitous class of small regulatory RNAs that serve as posttranscriptional regulators of gene expression. Previous work has suggested that HIV-1 might subvert the function of the cellular miRNA machinery by expressing viral miRNAs or by dramatically altering the level of cellular miRNA expression. Using very sensitive approaches, we now demonstrate that neither of these ideas is in fact correct. Moreover, HIV-1 transcripts appear to largely avoid regulation by cellular miRNAs by adopting an extensive RNA secondary structure that occludes the ability of cellular miRNAs to interact with viral mRNAs. Together, these data suggest that HIV-1, rather than seeking to control miRNA function in infected cells, has instead evolved a mechanism to become largely invisible to cellular miRNA effector mechanisms.