Phosphorylation of octamer-binding transcriptional factor may be correlated with H2B histone gene repression during 12-O-tetradecanoylphorbol 13-acetate-dependent differentiation of HL-60 cells.
To gain insight on the role of transacting factors in the regulatory mechanism of H2B histone gene expression during the differentiation of HL-60 cells by 12-O-tetradecanoylphorbol 13-acetate (TPA), the binding pattern of nuclear proteins to various elements in the human H2B histone gene upstream region have been investigated with DNase I footprinting and DNA mobility shift assay. The level of H2B histone mRNA rapidly reduced at 24 h in TPA-treated HL-60 cells. The H2B histone mRNA was repressed in proportion to the concentration of TPA. In DNase I footprinting analysis, one nuclear factor (octamer-binding transcription factor, OTF) bound at -42 bp (octamer motif), before and after TPA-induced differentiation of HL-60 cells. One DNA-protein complex (OTF) was formed by DNA mobility shift assay when octamer element was incubated with nuclear extract of undifferentiated HL-60 cells. In DNA mobilith shift assay, OTF vanished, and phosphorylated OTF (p-OTF) newly appeared during TPA-induced differentiation. p-OTF was not detected after pretreatment of the protein kinase C inhibitor, staurosporin, but was not changed after CHX treatment. TPA-induced repression of H2B histone mRNA was also restored after pretreatment of staurosporin. These results suggest that OTF is phosphorylated by protein kinase C during TPA-induced differentiation of HL-60 and the transcriptional repression of the H2B histone gene may be mediated by protein kinase C-dependent phosphorylation of OTF.
Lim
Kyu
K
Department of Biochemistry, College of Medicine, Chungnam National University, Jungu, Daejeon 301-747, Korea. kyulim@cnu.ac.kr
Kang
Yong-Sun
YS
Son
Mee-Young
MY
Park
Chung
C
Yun
Eun-Jin
EJ
Song
Kyoung-Sub
KS
Kim
Jong-Seok
JS
Kim
Young-Rae
YR
Park
Jong-Il
JI
Yoon
Wan-Hee
WH
Park
Seung-Kiel
SK
Hwang
Byung-Doo
BD
D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000187:drug effects* / Q000235:genetics; D004305:Dose-Response Relationship, Drug; D024202:Electrophoretic Mobility Shift Assay; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D018922:HL-60 Cells; D006657:Histones / Q000235:genetics*; D006801:Humans; D009687:Nuclear Proteins / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
D006657:Histones; D009687:Nuclear Proteins; D009841:Oligonucleotides; D010703:Phorbol Esters; D012333:RNA, Messenger; D014157:Transcription Factors