Upstream stimulatory factors stimulate transcription through E-box motifs in the PF4 gene in megakaryocytes.
Platelet factor 4 (PF4) is expressed during megakaryocytic differentiation. We previously demonstrated that the homeodomain proteins (myeloid ecotropic integration site 1 [MEIS1], Pbx-regulating protein 1 [PREP1], and pre-B-cell leukemia transcription factors [PBXs]) bind to the novel regulatory element tandem repeat of MEIS1 binding element [TME] and transactivate the rat PF4 promoter. In the present study, we investigated and identified other TME binding proteins in megakaryocytic HEL cells using mass spectrometry. Among identified proteins, we focused on upstream stimulatory factor (USF1) and USF2 and investigated their effects on the PF4 promoter. USF1 and 2 bound to the E-box motif in the TME and strongly transactivated the PF4 promoter. Furthermore, physiologic bindings of USF1 and 2 to the TME in rat megakaryocytes were demonstrated by the chromatin immunoprecipitation (ChIP) assay. Interestingly, the E-box motif in the TME was conserved in TME-like sequences of both the human and mouse PF4 promoters. USF1 and 2 also bound to the human TME-like sequence and transactivated the human PF4 promoter. Expressions of USF1 and 2 were detected by reverse-transcriptase-polymerase chain reaction (RT-PCR) in the human megakaryocytes derived from CD34+ cells. Thus, these studies demonstrate that the novel TME binding transcription factors, USF1 and 2, transactivate rat and human PF4 promoters and may play an important role in megakaryocytic gene expression.
Okada
Yoshiaki
Y
Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka, Japan.
Matsuura
Eri
E
Tozuka
Zenzaburo
Z
Nagai
Ryohei
R
Watanabe
Ayako
A
Matsumoto
Kayoko
K
Yasui
Kazuta
K
Jackman
Robert W
RW
Nakano
Toru
T
Doi
Takefumi
T
D020816:Amino Acid Motifs; D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002454:Cell Differentiation; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D013058:Mass Spectrometry; D008533:Megakaryocytes / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010957:Plasmids / Q000378:metabolism; D010978:Platelet Factor 4 / Q000737:chemistry* / Q000378:metabolism; D011233:Precipitin Tests; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection; D051720:Upstream Stimulatory Factors
D002843:Chromatin; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D014157:Transcription Factors; C497181:USF1 protein, human; C497184:USF2 protein, human; D051720:Upstream Stimulatory Factors; C497183:Usf1 protein, mouse; C497182:Usf1 protein, rat; C497186:Usf2 protein, mouse; C497187:Usf2 protein, rat; D010978:Platelet Factor 4