Purification and functional analysis of a novel leucine-zipper/nucleotide-fold protein, BZAP45, stimulating cell cycle regulated histone H4 gene transcription

Regulation of histone gene transcription at the G1/S phase transition via t he Site II cell cycle control element is distinct from E2F-dependent mechan isms operative at the growth factor-related restriction point. E2F-independ ent activation of historic H4 gene expression combines contributions of sev eral promoter factors, including HiNF-M/IRF2 and the HiNF-D/CDP-cut complex which contains pRB, CDK1, and cyclin A as non-DNA binding subunits. Mutati onal analyses suggest additional rate-limiting factors for Site II function . Using sequence-specific Site II DNA affinity chromatography, we identifie d a 45 kDa protein (KIAA0005 or BZAP45) that is embryonically expressed and phylogenetically conserved. Based on amino acid sequence analysis, BZAP45 contains a unique decapeptide that is part of a putative leucine-zipper pro tein with a nucleotide (ATP or GTP) binding fold. Bacterial expression of a full-length cDNA produces a 45 kDa protein. Binding studies reveal that hi ghly purified BZAP45 does not interact with Site II, suggesting that BZAP45 function may require partner proteins. Forced expression of BZAP45 strongl y stimulates H4 promoter (nt -215 to -1)/CAT reporter gene activity. Deleti on analyses and point mutations indicate that BZAP45 enhances H4 gene trans cription through Site II. Thus, BZAP45 is a novel regulatory factor that co ntributes to transcriptional control at the GUS phase transition.