Transcriptional element H4-site II of cell cycle regulated human H4 histone genes is a multipartite protein/DNA interaction site for factors HiNF-D, HiNF-M, and HiNF-P: involvement of phosphorylation.
Cell cycle regulated gene expression was studied by analyzing protein/DNA interactions occurring at the H4-Site II transcriptional element of H4 histone genes using several approaches. We show that this key proximal promoter element interacts with at least three distinct sequence-specific DNA binding activities, designated HiNF-D, HiNF-M, and HiNF-P. HiNF-D binds to an extended series of nucleotides, whereas HiNF-M and HiNF-P recognize sequences internal to the HiNF-D binding domain. Gel retardation assays show that HiNF-D and HiNF-M each are represented by two distinct protein/DNA complexes involving the same DNA binding activity. These results suggest that these factors are subject to post-translational modifications. Dephosphorylation experiments in vitro suggest that both electrophoretic mobility and DNA binding activities of HiNF-D and HiNF-M are sensitive to phosphatase activity. We deduce that these factors may require a basal level of phosphorylation for sequence specific binding to H4-Site II and may represent phosphoproteins occurring in putative hyper- and hypo-phosphorylated forms. Based on dramatic fluctuations in the ratio of the two distinct HiNF-D species both during hepatic development and the cell cycle in normal diploid cells, we postulate that this modification of HiNF-D is related to the cell cycle. However, in several tumor-derived and transformed cell types the putative hyperphosphorylated form of HiNF-D is constitutively present. These data suggest that deregulation of a phosphatase-sensitive post-translational modification required for HiNF-D binding is a molecular event that reflects abrogation of a mechanism controlling cell proliferation. Thus, phosphorylation and dephosphorylation of histone promoter factors may provide a basis for modulation of protein/DNA interactions and H4 histone gene transcription during the cell cycle and at the onset of quiescence and differentiation.
van Wijnen
A J
AJ
Department of Cell Biology, University of Massachusetts Medical School, Worcester 01655.
Ramsey-Ewing
A L
AL
Bortell
R
R
Owen
T A
TA
Lian
J B
JB
Stein
J L
JL
Stein
G S
GS
D001483:Base Sequence; D001665:Binding Sites; D002453:Cell Cycle; D002846:Chromatography, Affinity; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D006657:Histones / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids; D011499:Protein Processing, Post-Translational; D014157:Transcription Factors / Q000235:genetics*; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; C059062:H4TF-1 protein, human; C071109:HINF-D protein, human; D006657:Histones; D014157:Transcription Factors; D010749:Phosphoprotein Phosphatases