>POTED|NM_174981|3'UTR
   1 GTACAGTGGACAGCTTAGG

ID	Duplex structure	Position	Score	MFE
1	miRNA 3' aaaccagugugGUCAGGUGUaa 5' |||| ||| Target 5' --------gtaCAGTGGACAgc 3'	1 - 14	71.00	-5.30

Mutant ID	Mutant Position	Mutant Source
COSN31961891	9	COSMIC
COSN5028327	20	COSMIC

Mutant ID	Mutant Position	Mutant Source
rs1276157386	6	dbSNP
rs768070926	30	dbSNP
rs1296859362	36	dbSNP
rs776163294	38	dbSNP
rs1257431422	43	dbSNP
rs761656523	44	dbSNP
rs1160839455	50	dbSNP
rs1348516768	52	dbSNP
rs765204623	53	dbSNP
rs1384921280	57	dbSNP
rs201280833	59	dbSNP
rs1188473127	64	dbSNP
rs562566026	70	dbSNP
rs1444251120	83	dbSNP
rs1259775024	86	dbSNP
rs1204596512	99	dbSNP
rs1253985332	101	dbSNP
rs1287998387	102	dbSNP
rs531439387	107	dbSNP
rs1358455307	124	dbSNP
rs949601332	127	dbSNP
rs551550729	134	dbSNP
rs1246504868	163	dbSNP
rs1046652159	173	dbSNP
rs1317025735	188	dbSNP
rs1400018276	193	dbSNP
rs1309534072	209	dbSNP
rs1361805969	223	dbSNP
rs1300583192	235	dbSNP
rs1382358145	244	dbSNP
rs1454503773	271	dbSNP
rs1164152749	328	dbSNP
rs1473917905	336	dbSNP
rs1371479676	355	dbSNP
rs902653376	386	dbSNP
rs1162673308	387	dbSNP
rs1054400389	390	dbSNP
rs999679440	390	dbSNP
rs10222155	391	dbSNP
rs1490139468	410	dbSNP
rs1290357255	411	dbSNP
rs1211371276	412	dbSNP
rs1337055421	416	dbSNP
rs2849161	420	dbSNP
rs1269214726	436	dbSNP
rs368712337	457	dbSNP
rs2150359	506	dbSNP
rs1218780873	545	dbSNP
rs1341801209	564	dbSNP
rs1280700450	582	dbSNP
rs1403657757	608	dbSNP
rs1242841111	618	dbSNP
rs1349766853	630	dbSNP
rs893546574	640	dbSNP
rs1407378883	641	dbSNP
rs1007058464	648	dbSNP
rs1017913935	668	dbSNP
rs1388744034	675	dbSNP
rs1466246669	684	dbSNP
rs1425845488	710	dbSNP
rs1197305938	714	dbSNP
rs1407014733	725	dbSNP
rs1200806440	726	dbSNP
rs965058467	726	dbSNP
rs1435860863	731	dbSNP
rs1324174802	732	dbSNP
rs1375851641	736	dbSNP
rs1444250033	740	dbSNP
rs1222300474	758	dbSNP
rs772964507	761	dbSNP
rs998073102	763	dbSNP
rs1377100233	764	dbSNP
rs2334592	783	dbSNP
rs1439135868	785	dbSNP
rs1243628689	794	dbSNP
rs1334321449	797	dbSNP
rs4524196	835	dbSNP
rs8129783	858	dbSNP
rs4416309	860	dbSNP
rs1263161653	862	dbSNP
rs1402678495	864	dbSNP
rs1174376379	872	dbSNP
rs1413963989	878	dbSNP
rs1425868306	896	dbSNP
rs1174216026	904	dbSNP
rs1357895101	920	dbSNP
rs1212128432	931	dbSNP
rs1178975285	935	dbSNP
rs1458814884	936	dbSNP
rs1274298360	951	dbSNP
rs1202555058	952	dbSNP
rs1312264954	959	dbSNP
rs2183548	963	dbSNP
rs1457733388	967	dbSNP
rs1221416936	969	dbSNP
rs1177086239	970	dbSNP
rs1240424558	972	dbSNP
rs571254196	973	dbSNP
rs4490281	974	dbSNP
rs1379468310	978	dbSNP
rs1334962451	990	dbSNP
rs2150360	993	dbSNP
rs1473847384	999	dbSNP
rs1336192712	1028	dbSNP
rs1398911814	1040	dbSNP
rs1379534804	1074	dbSNP
rs1159034077	1090	dbSNP
rs141924343	1100	dbSNP
rs775908656	1114	dbSNP
rs879112925	1154	dbSNP
rs112360650	1158	dbSNP
rs1441304972	1170	dbSNP
rs1383074444	1172	dbSNP
rs2104806	1173	dbSNP
rs951399480	1181	dbSNP
rs983968509	1212	dbSNP
rs1333050297	1213	dbSNP
rs1017215658	1217	dbSNP
rs1487597321	1224	dbSNP
rs963461458	1237	dbSNP
rs1310198424	1245	dbSNP
rs990869065	1251	dbSNP
rs1341915934	1253	dbSNP
rs1356257564	1274	dbSNP
rs1313252075	1284	dbSNP
rs1244828107	1292	dbSNP
rs916558602	1332	dbSNP
rs949402531	1346	dbSNP
rs1428648704	1352	dbSNP
rs1359883707	1418	dbSNP
rs1298733569	1488	dbSNP
rs1421334281	1529	dbSNP
rs982108833	1564	dbSNP
rs1197363388	1574	dbSNP
rs1424064698	1577	dbSNP
rs1415429729	1578	dbSNP
rs1184272695	1598	dbSNP
rs1260396498	1607	dbSNP
rs796573791	1636	dbSNP
rs182956641	1655	dbSNP
rs201172569	1665	dbSNP
rs1481189971	1673	dbSNP
rs1217960248	1700	dbSNP
rs4471970	1703	dbSNP
rs1489527835	1716	dbSNP
rs1264082507	1721	dbSNP
rs1200071645	1725	dbSNP
rs935614341	1726	dbSNP
rs4555432	1734	dbSNP
rs202036840	1749	dbSNP
rs1292271108	1757	dbSNP
rs1212953767	1784	dbSNP
rs533807683	1795	dbSNP
rs760953377	1799	dbSNP
rs187301606	1801	dbSNP
rs200346536	1802	dbSNP
rs1401796928	1810	dbSNP
rs4239828	1835	dbSNP
rs4239829	1840	dbSNP
rs1423025103	1845	dbSNP
rs1476793180	1850	dbSNP
rs1351319053	1865	dbSNP
rs1172780521	1877	dbSNP
rs1464692979	1878	dbSNP
rs1373408393	1901	dbSNP
rs1423745426	1917	dbSNP
rs1195160057	1958	dbSNP
rs371500066	1961	dbSNP
rs1479089744	1965	dbSNP
rs1465315715	1966	dbSNP
rs1191034911	2000	dbSNP
rs1489006991	2028	dbSNP
rs8131141	2034	dbSNP
rs1300894860	2035	dbSNP
rs1403813877	2061	dbSNP
rs1196964279	2066	dbSNP
rs1415030391	2071	dbSNP
rs1258835261	2077	dbSNP
rs1220477818	2080	dbSNP
rs1310383805	2096	dbSNP
rs1300487694	2097	dbSNP
rs1297022601	2101	dbSNP
rs1369436455	2110	dbSNP
rs1337856446	2119	dbSNP
rs548047664	2123	dbSNP
rs1053922808	2127	dbSNP
rs142359277	2132	dbSNP
rs1172206780	2148	dbSNP
rs1308624578	2160	dbSNP
rs1376498040	2167	dbSNP
rs1175941710	2168	dbSNP
rs1434172698	2169	dbSNP
rs367816011	2178	dbSNP
rs567887040	2180	dbSNP
rs1269945186	2184	dbSNP
rs753979750	2197	dbSNP
rs893967934	2198	dbSNP
rs1457489301	2245	dbSNP
rs1238962120	2246	dbSNP
rs1200667404	2250	dbSNP
rs1482315467	2265	dbSNP
rs1255391039	2287	dbSNP
rs1233907668	2292	dbSNP
rs1313840735	2321	dbSNP
rs202146573	2324	dbSNP
rs1306811106	2325	dbSNP
rs1239083750	2337	dbSNP
rs1377253625	2338	dbSNP
rs1332942287	2348	dbSNP
rs1449879245	2350	dbSNP
rs1400847424	2385	dbSNP
rs1302872765	2391	dbSNP
rs1467032935	2399	dbSNP
rs1379814469	2419	dbSNP
rs1157246582	2426	dbSNP
rs1232941184	2435	dbSNP
rs1254634748	2447	dbSNP
rs1484326727	2455	dbSNP
rs887077189	2457	dbSNP
rs3017190	2467	dbSNP
rs1381224885	2473	dbSNP
rs1180712087	2482	dbSNP
rs1440858806	2493	dbSNP
rs1211939268	2500	dbSNP
rs112139263	2507	dbSNP
rs4046835	2508	dbSNP
rs1201040408	2509	dbSNP
rs1484200600	2514	dbSNP
rs1283778396	2534	dbSNP
rs1215706356	2535	dbSNP
rs73148745	2543	dbSNP
rs1311066646	2544	dbSNP
rs4046837	2582	dbSNP
rs1242193914	2585	dbSNP
rs1339472434	2586	dbSNP
rs1312818415	2603	dbSNP
rs545456990	2613	dbSNP
rs3017191	2621	dbSNP
rs1262525751	2633	dbSNP
rs1377859606	2642	dbSNP
rs1487377364	2643	dbSNP
rs1419524380	2662	dbSNP
rs1361596178	2663	dbSNP
rs3017192	2664	dbSNP
rs1195960445	2666	dbSNP
rs1262291709	2680	dbSNP
rs373410283	2681	dbSNP
rs1471934659	2705	dbSNP
rs191745103	2729	dbSNP
rs1368056246	2741	dbSNP
rs1188399378	2750	dbSNP
rs1432526699	2754	dbSNP
rs1192821993	2756	dbSNP
rs1488550298	2760	dbSNP
rs1287940263	2764	dbSNP
rs4046840	2785	dbSNP
rs1488114905	2793	dbSNP
rs1207779058	2797	dbSNP
rs1355229489	2808	dbSNP
rs1265651703	2842	dbSNP
rs1246437545	2847	dbSNP
rs1194152319	2854	dbSNP
rs2334594	2873	dbSNP
rs536821056	2874	dbSNP
rs2334595	2881	dbSNP
rs1399965081	2893	dbSNP
rs4046841	2897	dbSNP
rs1379207127	2915	dbSNP
rs1320707721	2929	dbSNP
rs1403937533	2931	dbSNP
rs1390370111	2939	dbSNP
rs556490059	2941	dbSNP
rs1164016252	2942	dbSNP
rs1463120790	2943	dbSNP
rs2334596	2944	dbSNP
rs1421608201	2949	dbSNP
rs1192993663	2950	dbSNP
rs1478111965	2954	dbSNP
rs1265177740	2962	dbSNP
rs2334597	2965	dbSNP
rs2334598	2967	dbSNP
rs757229848	2968	dbSNP
rs765781265	2969	dbSNP
rs1267665339	2992	dbSNP
rs1211243440	2993	dbSNP
rs576434152	2998	dbSNP
rs538743581	3003	dbSNP
rs1256872693	3020	dbSNP
rs1157722765	3032	dbSNP
rs183958834	3038	dbSNP
rs1358054851	3043	dbSNP
rs1464660555	3060	dbSNP
rs2334599	3082	dbSNP
rs4046842	3083	dbSNP
rs2334600	3086	dbSNP
rs1288978119	3121	dbSNP
rs879006654	3141	dbSNP
rs78806737	3145	dbSNP
rs1814006	3151	dbSNP
rs1443424049	3158	dbSNP
rs1299626984	3159	dbSNP
rs1814007	3164	dbSNP
rs2334601	3167	dbSNP
rs2334602	3168	dbSNP
rs758820384	3192	dbSNP
rs1466199480	3193	dbSNP
rs1421647904	3200	dbSNP
rs1277959134	3203	dbSNP
rs1477604202	3222	dbSNP
rs1378013573	3227	dbSNP
rs1200648734	3228	dbSNP
rs1402108262	3231	dbSNP
rs4046830	3234	dbSNP
rs1482369890	3239	dbSNP
rs1456180007	3240	dbSNP
rs1200136939	3241	dbSNP
rs141566892	3242	dbSNP
rs1262513931	3259	dbSNP
rs1430440668	3267	dbSNP
rs942402576	3269	dbSNP
rs113348606	3270	dbSNP
rs1455109194	3272	dbSNP
rs3080117	3276	dbSNP
rs1275818835	3278	dbSNP
rs1236883070	3279	dbSNP
rs1375035740	3279	dbSNP
rs1175869468	3281	dbSNP
rs28888521	3297	dbSNP
rs1447663913	3304	dbSNP
rs4098544	3312	dbSNP
rs1332223160	3319	dbSNP
rs900951037	3323	dbSNP
rs1425711532	3336	dbSNP
rs1175345229	3339	dbSNP
rs997834029	3345	dbSNP
rs1379340577	3350	dbSNP
rs28782309	3352	dbSNP
rs1178838435	3358	dbSNP
rs1470930238	3365	dbSNP
rs1235272287	3386	dbSNP
rs1208740319	3387	dbSNP
rs1442366346	3389	dbSNP
rs1378058438	3393	dbSNP
rs1421145415	3411	dbSNP
rs1159125203	3414	dbSNP
rs1348855172	3417	dbSNP
rs1351924348	3421	dbSNP
rs1308866648	3424	dbSNP
rs1241180464	3426	dbSNP
rs1375753032	3429	dbSNP
rs879066571	3434	dbSNP
rs879040057	3436	dbSNP
rs4098543	3439	dbSNP
rs1447368792	3448	dbSNP
rs1359739503	3457	dbSNP
rs8129761	3459	dbSNP
rs572090218	3461	dbSNP
rs1457839329	3462	dbSNP
rs1417682303	3465	dbSNP
rs8131497	3466	dbSNP
rs1158980709	3475	dbSNP
rs1402092768	3476	dbSNP
rs1373855839	3477	dbSNP
rs1180838160	3478	dbSNP
rs1258876828	3481	dbSNP
rs1472840317	3481	dbSNP
rs554002512	3482	dbSNP
rs573864437	3487	dbSNP
rs1308993732	3489	dbSNP
rs1487567689	3490	dbSNP
rs1025771550	3502	dbSNP
rs1285793171	3505	dbSNP
rs3017193	3514	dbSNP
rs542762558	3519	dbSNP
rs1217729342	3524	dbSNP
rs1319012138	3526	dbSNP
rs1374871135	3537	dbSNP
rs149752065	3538	dbSNP
rs8131522	3540	dbSNP
rs2878225	3541	dbSNP
rs1285034300	3546	dbSNP
rs1398101897	3548	dbSNP
rs545110258	3549	dbSNP
rs1298600759	3551	dbSNP
rs1219257984	3560	dbSNP
rs1267096614	3561	dbSNP
rs1347831661	3565	dbSNP
rs1468887206	3566	dbSNP
rs1405495825	3567	dbSNP
rs3080118	3575	dbSNP
rs755846239	3575	dbSNP
rs1419418348	3585	dbSNP
rs565000593	3589	dbSNP
rs1477131776	3592	dbSNP
rs527252161	3594	dbSNP
rs1236150726	3615	dbSNP
rs1441144654	3617	dbSNP
rs887023714	3618	dbSNP
rs1005889921	3619	dbSNP
rs1221134414	3621	dbSNP
rs1335667124	3629	dbSNP
rs1177493948	3634	dbSNP
rs1216817299	3638	dbSNP
rs1317286694	3639	dbSNP
rs1278763656	3640	dbSNP
rs1383751923	3644	dbSNP
rs754658600	3648	dbSNP
rs1292683181	3651	dbSNP
rs4046831	3655	dbSNP
rs113198593	3660	dbSNP
rs1164983571	3663	dbSNP
rs1016730287	3678	dbSNP
rs1421491631	3683	dbSNP
rs1166332311	3687	dbSNP
rs547092366	3692	dbSNP
rs1464644203	3696	dbSNP
rs567907733	3701	dbSNP
rs1397240266	3703	dbSNP
rs1171707513	3705	dbSNP
rs1430758814	3710	dbSNP
rs1423540851	3712	dbSNP
rs1190905310	3719	dbSNP
rs8130064	3720	dbSNP
rs1462438294	3721	dbSNP
rs1331079709	3725	dbSNP
rs2334603	3726	dbSNP
rs2334604	3734	dbSNP
rs2334605	3739	dbSNP
rs1444592744	3741	dbSNP
rs958806222	3742	dbSNP
rs1280404991	3745	dbSNP
rs1234681282	3748	dbSNP
rs550225230	3748	dbSNP
rs1372807122	3752	dbSNP
rs2334606	3759	dbSNP
rs1378753917	3761	dbSNP
rs1226643873	3763	dbSNP
rs1295210452	3768	dbSNP
rs1445463295	3771	dbSNP
rs1335334346	3772	dbSNP
rs1218001081	3785	dbSNP
rs550497058	3787	dbSNP
rs1357871452	3793	dbSNP
rs1175868825	3802	dbSNP
rs1454981326	3804	dbSNP
rs1023663831	3810	dbSNP
rs2334607	3816	dbSNP
rs1204751501	3820	dbSNP
rs1399081523	3824	dbSNP
rs372760414	3826	dbSNP
rs1482073827	3827	dbSNP
rs1255334771	3835	dbSNP
rs982068009	3839	dbSNP
rs868756316	3842	dbSNP
rs1306745960	3849	dbSNP
rs1475815759	3851	dbSNP
rs1335908008	3854	dbSNP
rs2334608	3858	dbSNP
rs4263202	3868	dbSNP
rs1170643603	3869	dbSNP
rs2334609	3870	dbSNP
rs569903787	3879	dbSNP
rs372059940	3886	dbSNP
rs1330008685	3889	dbSNP
rs4046832	3892	dbSNP
rs1358736745	3906	dbSNP
rs8130373	3919	dbSNP
rs1314822254	3922	dbSNP
rs1419337098	3926	dbSNP
rs1356252454	3930	dbSNP
rs1364543021	3938	dbSNP
rs1184149936	3944	dbSNP
rs9967975	3945	dbSNP
rs4046833	3953	dbSNP
rs1422098843	3957	dbSNP
rs1255445071	3958	dbSNP
rs538878542	3969	dbSNP
rs1486588419	3973	dbSNP
rs1276284137	3975	dbSNP
rs4046834	3976	dbSNP
rs1214871650	3982	dbSNP
rs558603959	3983	dbSNP
rs1242035232	3987	dbSNP
rs1310060260	3988	dbSNP
rs1293320108	3997	dbSNP
rs1212656065	3998	dbSNP
rs1257561778	4010	dbSNP
rs1357839927	4011	dbSNP
rs1296752492	4013	dbSNP
rs8127303	4015	dbSNP
rs1381997653	4019	dbSNP
rs1385996068	4026	dbSNP
rs962804845	4031	dbSNP
rs1487702604	4032	dbSNP
rs1160488903	4036	dbSNP
rs1190900851	4038	dbSNP
rs142965563	4040	dbSNP
rs201948634	4043	dbSNP
rs4046827	4056	dbSNP
rs1476152492	4066	dbSNP
rs529847566	4070	dbSNP
rs1260915443	4072	dbSNP
rs1192753340	4074	dbSNP
rs565856598	4080	dbSNP
rs1447336016	4081	dbSNP
rs1263472890	4089	dbSNP
rs1424983400	4092	dbSNP
rs1355166071	4094	dbSNP
rs768912110	4098	dbSNP
rs2334610	4099	dbSNP
rs1273273773	4101	dbSNP
rs923943517	4102	dbSNP
rs1337659736	4103	dbSNP
rs1276855613	4110	dbSNP
rs1227215187	4111	dbSNP
rs1412978392	4134	dbSNP
rs4263203	4135	dbSNP
rs1376123991	4136	dbSNP
rs3999506	4138	dbSNP
rs367675423	4141	dbSNP
rs1393946467	4158	dbSNP
rs1326730471	4164	dbSNP
rs1463072284	4173	dbSNP
rs1296423482	4185	dbSNP
rs8130773	4189	dbSNP
rs1169846645	4195	dbSNP
rs935556454	4196	dbSNP
rs1374616562	4197	dbSNP
rs1196405326	4206	dbSNP
rs1450702817	4208	dbSNP
rs1253229341	4215	dbSNP
rs1328070310	4227	dbSNP
rs1456050541	4228	dbSNP
rs1256812533	4232	dbSNP
rs1195760558	4235	dbSNP
rs1272302433	4245	dbSNP
rs1231509694	4252	dbSNP
rs9967972	4253	dbSNP

- Nature methods, 2011

Cross-linking and immunoprecipitation (CLIP) is increasingly used to map transcriptome-wide binding sites of RNA-binding proteins. We developed a method for CLIP data analysis, and applied it to compare CLIP with photoactivatable ribonucleoside-enhanced CLIP (PAR-CLIP) and to uncover how differences in cross-linking and ribonuclease digestion affect the identified sites. We found only small differences in accuracies of these methods in identifying binding sites of HuR, which binds low-complexity sequences, and Argonaute 2, which has a complex binding specificity. We found that cross-link-induced mutations led to single-nucleotide resolution for both PAR-CLIP and CLIP. Our results confirm the expectation from original CLIP publications that RNA-binding proteins do not protect their binding sites sufficiently under the denaturing conditions used during the CLIP procedure, and we show that extensive digestion with sequence-specific RNases strongly biases the recovered binding sites. This bias can be substantially reduced by milder nuclease digestion conditions.

- Genes & development, 2011

MicroRNAs are important regulators in many cellular processes, including stem cell self-renewal. Recent studies demonstrated their function as pluripotency factors with the capacity for somatic cell reprogramming. However, their role in human embryonic stem (ES) cells (hESCs) remains poorly understood, partially due to the lack of genome-wide strategies to identify their targets. Here, we performed comprehensive microRNA profiling in hESCs and in purified neural and mesenchymal derivatives. Using a combination of AGO cross-linking and microRNA perturbation experiments, together with computational prediction, we identified the targets of the miR-302/367 cluster, the most abundant microRNAs in hESCs. Functional studies identified novel roles of miR-302/367 in maintaining pluripotency and regulating hESC differentiation. We show that in addition to its role in TGF-beta signaling, miR-302/367 promotes bone morphogenetic protein (BMP) signaling by targeting BMP inhibitors TOB2, DAZAP2, and SLAIN1. This study broadens our understanding of microRNA function in hESCs and is a valuable resource for future studies in this area.

- Nature, 2013

Circular RNAs (circRNAs) in animals are an enigmatic class of RNA with unknown function. To explore circRNAs systematically, we sequenced and computationally analysed human, mouse and nematode RNA. We detected thousands of well-expressed, stable circRNAs, often showing tissue/developmental-stage-specific expression. Sequence analysis indicated important regulatory functions for circRNAs. We found that a human circRNA, antisense to the cerebellar degeneration-related protein 1 transcript (CDR1as), is densely bound by microRNA (miRNA) effector complexes and harbours 63 conserved binding sites for the ancient miRNA miR-7. Further analyses indicated that CDR1as functions to bind miR-7 in neuronal tissues. Human CDR1as expression in zebrafish impaired midbrain development, similar to knocking down miR-7, suggesting that CDR1as is a miRNA antagonist with a miRNA-binding capacity ten times higher than any other known transcript. Together, our data provide evidence that circRNAs form a large class of post-transcriptional regulators. Numerous circRNAs form by head-to-tail splicing of exons, suggesting previously unrecognized regulatory potential of coding sequences.

- 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.

- Genome biology, 2014

BACKGROUND: Various microRNAs (miRNAs) are up- or downregulated in tumors. However, the repression of cognate miRNA targets responsible for the phenotypic effects of this dysregulation in patients remains largely unexplored. To define miRNA targets and associated pathways, together with their relationship to outcome in breast cancer, we integrated patient-paired miRNA-mRNA expression data with a set of validated miRNA targets and pathway inference. RESULTS: To generate a biochemically-validated set of miRNA-binding sites, we performed argonaute-2 photoactivatable-ribonucleoside-enhanced crosslinking and immunoprecipitation (AGO2-PAR-CLIP) in MCF7 cells. We then defined putative miRNA-target interactions using a computational model, which ranked and selected additional TargetScan-predicted interactions based on features of our AGO2-PAR-CLIP binding-site data. We subselected modeled interactions according to the abundance of their constituent miRNA and mRNA transcripts in tumors, and we took advantage of the variability of miRNA expression within molecular subtypes to detect miRNA repression. Interestingly, our data suggest that miRNA families control subtype-specific pathways; for example, miR-17, miR-19a, miR-25, and miR-200b show high miRNA regulatory activity in the triple-negative, basal-like subtype, whereas miR-22 and miR-24 do so in the HER2 subtype. An independent dataset validated our findings for miR-17 and miR-25, and showed a correlation between the expression levels of miR-182 targets and overall patient survival. Pathway analysis associated miR-17, miR-19a, and miR-200b with leukocyte transendothelial migration. CONCLUSIONS: We combined PAR-CLIP data with patient expression data to predict regulatory miRNAs, revealing potential therapeutic targets and prognostic markers in breast cancer.