title: Upstream stimulatory factors stimulate transcription through E-box motifs in the PF4 gene in megakaryocytes.
abstract: 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.
authors.affiliation: Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka, Japan.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Okada
authors.forename: Eri
authors.initials: E
authors.lastname: Matsuura
authors.forename: Zenzaburo
authors.initials: Z
authors.lastname: Tozuka
authors.forename: Ryohei
authors.initials: R
authors.lastname: Nagai
authors.forename: Ayako
authors.initials: A
authors.lastname: Watanabe
authors.forename: Kayoko
authors.initials: K
authors.lastname: Matsumoto
authors.forename: Kazuta
authors.initials: K
authors.lastname: Yasui
authors.forename: Robert W
authors.initials: RW
authors.lastname: Jackman
authors.forename: Toru
authors.initials: T
authors.lastname: Nakano
authors.forename: Takefumi
authors.initials: T
authors.lastname: Doi
mesh_terms: 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
chemical_list: 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