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Two proteins specifically binding to separate regions of the human histone H4 promoter were identified in nuclear extracts prepared from synchronized S-phase HeLa cells. Competition experiments with H4 promoter mutants and DNase protection assays ("footprinting") demonstrate that these factors bind to regions of the H4 promoter that are essential for maximal expression in vitro. One of these factors (H4TF-1) binds to sequences between -80 and -110 base pairs upstream of the H4 cap site, whereas the other (H4TF-2) binds to the H4 subtype-specific sequence element immediately upstream from the "TATA" homology. Neither of these activities can efficiently bind to any of the other histone gene subtypes or simian virus 40 DNA. Binding of H4TF-1 to the distal region of the pHu4A histone H4 promoter is inhibited competitively with varying efficiency by four of six human histone H4 genes cloned in this laboratory, whereas efficient competition for binding of H4TF-2 is exhibited by five of the six H4 genes. Since both of these factors bind to significant regions of the pHu4A histone H4 promoter and can be bound by several different human H4 genes, we believe that they are important for maximal transcription of the gene and that they may be involved in its regulated expression during the cell cycle.

Distinct transcription factors bind specifically to two regions of the human histone H4 promoter.