0 15 Transcriptional transcriptional JJ 16 26 activation activation NN 27 29 of of IN 30 35 human human JJ 36 40 zeta zeta NN 41 42 2 2 CD 43 49 globin globin NN 50 58 promoter promoter NN 59 61 by by IN 62 65 the the DT 66 71 alpha alpha NN 72 78 globin globin NN 79 89 regulatory regulatory JJ 90 97 element element NN 98 99 ( ( ( 99 104 HS-40 HS-40 NNP 104 105 ) ) ) 105 106 : : : 107 117 functional functional JJ 118 122 role role NN 123 125 of of IN 126 134 specific specific JJ 135 142 nuclear nuclear JJ 143 153 factor-DNA factor-dna NN 154 163 complexes complex NNS 163 164 . . . 166 168 We we PRP 169 176 studied study VBD 177 180 the the DT 181 191 functional functional JJ 192 203 interaction interaction NN 204 211 between between IN 212 217 human human JJ 218 227 embryonic embryonic JJ 228 232 zeta zeta NN 233 234 2 2 CD 235 241 globin globin NN 242 250 promoter promoter NN 251 254 and and CC 255 258 the the DT 259 264 alpha alpha NN 265 271 globin globin NN 272 282 regulatory regulatory JJ 283 290 element element NN 291 292 ( ( ( 292 297 HS-40 hs-40 NN 297 298 ) ) ) 299 306 located located JJ 307 309 40 40 CD 310 312 kb kb NN 313 321 upstream upstream RB 322 324 of of IN 325 328 the the DT 329 333 zeta zeta NN 334 335 2 2 CD 336 342 globin globin NN 343 347 gene gene NN 347 348 . . . 349 351 It it PRP 352 355 was be VBD 356 361 shown show VBN 362 364 by by IN 365 374 transient transient JJ 375 385 expression expression NN 386 391 assay assay NN 392 396 that that IN 397 402 HS-40 HS-40 NNP 403 410 behaved behave VBD 411 413 as as IN 414 416 an an DT 417 426 authentic authentic JJ 427 435 enhancer enhancer NN 436 439 for for IN 440 450 high-level high-level JJ 451 455 zeta zeta NN 456 457 2 2 CD 458 464 globin globin NN 465 473 promoter promoter NN 474 482 activity activity NN 483 485 in in IN 486 490 K562 k562 NN 491 496 cells cell NNS 496 497 , , , 498 500 an an DT 501 510 erythroid erythroid JJ 511 515 cell cell NN 516 520 line line NN 521 523 of of IN 524 533 embryonic embryonic JJ 534 540 and/or and/or CC 541 546 fetal fetal JJ 547 553 origin origin NN 553 554 . . . 555 563 Although although IN 564 573 sequences sequence NNS 574 581 located located JJ 582 589 between between IN 590 594 -559 -559 CD 595 598 and and CC 599 602 -88 -88 CD 603 605 of of IN 606 609 the the DT 610 614 zeta zeta NN 615 616 2 2 CD 617 623 globin globin NN 624 628 gene gene NN 629 633 were be VBD 634 645 dispensable dispensable JJ 646 649 for for IN 650 653 its its PRP$ 654 664 expression expression NN 665 667 on on IN 668 680 enhancerless enhancerless JJ 681 689 plasmids plasmid NNS 689 690 , , , 691 695 they they PRP 696 700 were be VBD 701 709 required require VBN 710 713 for for IN 714 717 the the DT 718 723 HS-40 HS-40 NNP 724 741 enhancer-mediated enhancer-mediated JJ 742 750 activity activity NN 751 753 of of IN 754 757 the the DT 758 762 zeta zeta NN 763 764 2 2 CD 765 771 globin globin NN 772 780 promoter promoter NN 780 781 . . . 782 795 Site-directed site-directed JJ 796 807 mutagenesis mutagenesis NN 808 820 demonstrated demonstrate VBD 821 825 that that IN 826 830 this this DT 831 836 HS-40 hs-40 NN 837 850 enhancer-zeta enhancer-zeta NN 851 852 2 2 CD 853 859 globin globin NN 860 868 promoter promoter NN 869 880 interaction interaction NN 881 883 is be VBZ 884 892 mediated mediate VBN 893 895 by by IN 896 899 the the DT 900 903 two two CD 904 910 GATA-1 gata-1 NN 911 917 factor factor NN 918 925 binding binding NN 926 932 motifs motif NNS 933 940 located locate VBN 941 943 at at IN 944 948 -230 -230 CD 949 952 and and CC 953 957 -104 -104 CD 957 958 , , , 959 971 respectively respectively RB 971 972 . . . 973 976 The the DT 977 987 functional functional JJ 988 995 domains domain NNS 996 998 of of IN 999 1004 HS-40 HS-40 NNP 1005 1009 were be VBD 1010 1014 also also RB 1015 1021 mapped map VBN 1021 1022 . . . 1023 1026 Bal Bal NNP 1027 1029 31 31 CD 1030 1038 deletion deletion NN 1039 1046 mapping mapping NN 1047 1051 data datum NNS 1052 1061 suggested suggest VBD 1062 1066 that that IN 1067 1070 one one CD 1071 1077 GATA-1 gata-1 NN 1078 1083 motif motif NN 1083 1084 , , , 1085 1088 one one CD 1089 1091 GT gt NN 1092 1097 motif motif NN 1097 1098 , , , 1099 1102 and and CC 1103 1106 two two CD 1107 1116 NF-E2/AP1 nf-e2/ap1 NN 1117 1123 motifs motif NNS 1124 1132 together together RB 1133 1139 formed form VBD 1140 1143 the the DT 1144 1154 functional functional JJ 1155 1159 core core NN 1160 1162 of of IN 1163 1168 HS-40 hs-40 NN 1169 1171 in in IN 1172 1175 the the DT 1176 1194 erythroid-specific erythroid-specific JJ 1195 1205 activation activation NN 1206 1208 of of IN 1209 1212 the the DT 1213 1217 zeta zeta NN 1218 1219 2 2 CD 1220 1226 globin globin NN 1227 1235 promoter promoter NN 1235 1236 . . . 1237 1250 Site-directed site-directed JJ 1251 1262 mutagenesis mutagenesis NN 1263 1270 further further RBR 1271 1283 demonstrated demonstrate VBD 1284 1288 that that IN 1289 1292 the the DT 1293 1301 enhancer enhancer NN 1302 1310 function function NN 1311 1313 of of IN 1314 1317 one one CD 1318 1320 of of IN 1321 1324 the the DT 1325 1328 two two CD 1329 1338 NF-E2/AP1 nf-e2/ap1 NN 1339 1345 motifs motif NNS 1346 1348 of of IN 1349 1354 HS-40 HS-40 NNP 1355 1357 is be VBZ 1358 1366 mediated mediate VBN 1367 1374 through through IN 1375 1378 its its PRP$ 1379 1386 binding binding NN 1387 1389 to to TO 1390 1395 NF-E2 nf-e2 NN 1396 1399 but but CC 1400 1403 not not RB 1404 1407 AP1 ap1 NN 1408 1421 transcription transcription NN 1422 1428 factor factor NN 1428 1429 . . . 1430 1437 Finally finally RB 1437 1438 , , , 1439 1441 we we PRP 1442 1445 did do VBD 1446 1453 genomic genomic JJ 1454 1466 footprinting footprinting NN 1467 1469 of of IN 1470 1473 the the DT 1474 1479 HS-40 HS-40 NNP 1480 1488 enhancer enhancer NN 1489 1495 region region NN 1496 1498 in in IN 1499 1503 K562 k562 NN 1504 1509 cells cell NNS 1509 1510 , , , 1511 1516 adult adult JJ 1517 1526 nucleated nucleated JJ 1527 1540 erythroblasts erythroblast NNS 1540 1541 , , , 1542 1545 and and CC 1546 1555 different different JJ 1556 1568 nonerythroid nonerythroid JJ 1569 1574 cells cell NNS 1574 1575 . . . 1576 1579 All all DT 1580 1588 sequence sequence NN 1589 1595 motifs motif NNS 1596 1602 within within IN 1603 1606 the the DT 1607 1617 functional functional JJ 1618 1622 core core NN 1623 1625 of of IN 1626 1631 HS-40 HS-40 NNP 1631 1632 , , , 1633 1635 as as IN 1636 1642 mapped map VBN 1643 1645 by by IN 1646 1655 transient transient JJ 1656 1666 expression expression NN 1667 1675 analysis analysis NN 1675 1676 , , , 1677 1685 appeared appear VBD 1686 1688 to to TO 1689 1693 bind bind VB 1694 1695 a a DT 1696 1703 nuclear nuclear JJ 1704 1713 factor(s) factor(s) NN 1714 1716 in in IN 1717 1723 living live VBG 1724 1728 K562 k562 NN 1729 1734 cells cell NNS 1735 1738 but but CC 1739 1742 not not RB 1743 1745 in in IN 1746 1758 nonerythroid nonerythroid JJ 1759 1764 cells cell NNS 1764 1765 . . . 1766 1768 On on IN 1769 1772 the the DT 1773 1778 other other JJ 1779 1783 hand hand NN 1783 1784 , , , 1785 1789 only only RB 1790 1793 one one CD 1794 1796 of of IN 1797 1800 the the DT 1801 1811 apparently apparently RB 1812 1825 nonfunctional nonfunctional JJ 1826 1834 sequence sequence NN 1835 1841 motifs motif NNS 1842 1845 was be VBD 1846 1851 bound bind VBN 1852 1856 with with IN 1857 1864 factors factor NNS 1865 1867 in in FW 1868 1872 vivo vivo FW 1872 1873 . . . 1874 1876 In in IN 1877 1887 comparison comparison NN 1888 1890 to to TO 1891 1895 K562 k562 NN 1895 1896 , , , 1897 1906 nucleated nucleated JJ 1907 1920 erythroblasts erythroblast NNS 1921 1925 from from IN 1926 1931 adult adult JJ 1932 1937 human human JJ 1938 1942 bone bone NN 1943 1949 marrow marrow NN 1950 1959 exhibited exhibit VBD 1960 1961 a a DT 1962 1969 similar similar JJ 1970 1973 but but CC 1974 1986 nonidentical nonidentical JJ 1987 1994 pattern pattern NN 1995 1997 of of IN 1998 2005 nuclear nuclear JJ 2006 2012 factor factor NN 2013 2020 binding binding NN 2021 2023 in in FW 2024 2028 vivo vivo FW 2029 2031 at at IN 2032 2035 the the DT 2036 2041 HS-40 hs-40 NN 2042 2048 region region NN 2048 2049 . . . 2050 2055 These these DT 2056 2060 data datum NNS 2061 2068 suggest suggest VBP 2069 2073 that that IN 2074 2089 transcriptional transcriptional JJ 2090 2100 activation activation NN 2101 2103 of of IN 2104 2109 human human JJ 2110 2119 embryonic embryonic JJ 2120 2124 zeta zeta NN 2125 2126 2 2 CD 2127 2133 globin globin NN 2134 2138 gene gene NN 2139 2142 and and CC 2143 2146 the the DT 2147 2158 fetal/adult fetal/adult JJ 2159 2164 alpha alpha NN 2165 2171 globin globin NN 2172 2177 genes gene NNS 2178 2180 is be VBZ 2181 2189 mediated mediate VBN 2190 2192 by by IN 2193 2202 erythroid erythroid JJ 2203 2216 cell-specific cell-specific JJ 2217 2220 and and CC 2221 2234 developmental developmental JJ 2235 2249 stage-specific stage-specific JJ 2250 2257 nuclear nuclear JJ 2258 2268 factor-DNA factor-dna NN 2269 2278 complexes complex NNS 2279 2284 which which WDT 2285 2289 form form VBP 2290 2292 at at IN 2293 2296 the the DT 2297 2305 enhancer enhancer NN 2306 2307 ( ( ( 2307 2312 HS-40 HS-40 NNP 2312 2313 ) ) ) 2314 2317 and and CC 2318 2321 the the DT 2322 2328 globin globin NN 2329 2338 promoters promoter NNS 2338 2339 . . .