Sumoylation of p45/NF-E2: nuclear positioning and transcriptional activation of the mammalian beta-like globin gene locus.
NF-E2 is a transcription activator for the regulation of a number of erythroid- and megakaryocytic lineage-specific genes. Here we present evidence that the large subunit of mammalian NF-E2, p45, is sumoylated in vivo in human erythroid K562 cells and in mouse fetal liver. By in vitro sumoylation reaction and DNA transfection experiments, we show that the sumoylation occurs at lysine 368 (K368) of human p45/NF-E2. Furthermore, p45 sumoylation enhances the transactivation capability of NF-E2, and this is accompanied by an increase of the NF-E2 DNA binding affinity. More interestingly, we have found that in K562 cells, the beta-globin gene loci in the euchromatin regions are predominantly colocalized with the nuclear bodies promyelocytic leukemia protein (PML) oncogenic domains that are enriched with the PML, SUMO-1, RNA polymerase II, and sumoylatable p45/NF-E2. Chromatin immunoprecipitation assays further showed that the intact sumoylation site of p45/NF-E2 is required for its binding to the DNase I-hypersensitive sites of the beta-globin locus control region. Finally, we demonstrated by stable transfection assay that only the wild-type p45, but not its mutant form p45 (K368R), could efficiently rescue beta-globin gene expression in the p45-null, erythroid cell line CB3. These data together point to a model of mammalian beta-like globin gene activation by sumoylated p45/NF-E2 in erythroid cells.
Shyu
Yu-Chiau
YC
Institute of Molecular Biology, Academia Sinica, Nankang, Taipei 115, Taiwan, Republic of China.
Lee
Tung-Liang
TL
Ting
Chun-Yuan
CY
Wen
Shau-Ching
SC
Hsieh
Lie-Jiau
LJ
Li
Yueh-Chun
YC
Hwang
Jau-Lang
JL
Lin
Chyi-Chyang
CC
Shen
C-K James
CK
D021581:Active Transport, Cell Nucleus; D000818:Animals; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D041905:Erythroid Cells / Q000378:metabolism; D005914:Globins / Q000235:genetics*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D051379:Mice; D050984:NF-E2 Transcription Factor, p45 Subunit / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012319:RNA Polymerase II / Q000378:metabolism; D025842:SUMO-1 Protein / Q000378:metabolism*; D015533:Transcriptional Activation / Q000235:genetics*
D050984:NF-E2 Transcription Factor, p45 Subunit; C494692:NFE2 protein, human; D025842:SUMO-1 Protein; D005914:Globins; D004247:DNA; D012319:RNA Polymerase II; D008239:Lysine