Dailey L, Roberts S B, Heintz N
Laboratory of Molecular Biology, Rockefeller University, New York, New York 10021.
Genes Dev. 1988 Dec;2(12B):1700-12. doi: 10.1101/gad.2.12b.1700.
Transcription of the cell-cycle-regulated human histone genes increases approximately fivefold during S phase. One step toward the elucidation of the biochemical mechanisms that govern cell-cycle-regulated expression of these genes is to purify and characterize the transcription factors that regulate these promoters. Here, we describe the purification of two previously identified factors, H4TF-1 and H4TF-2, which bind the human histone H4 promoter. Purification was achieved through a combination of ion-exchange and oligonucleotide affinity chromatography. On the basis of analysis of purified fractions by SDS-polyacrylamide gels and UV cross-linking, we believe that H4TF-1 is two polypeptides of 105 and 110 kD. This factor binds to a GC-rich DNA sequence required for maximal expression of the H4 gene but does not bind to any Sp1 consensus elements tested. H4TF-2 is a 65-kD protein that binds specifically to sequences within that highly conserved H4 subtype-specific consensus promoter element. Both highly purified factors activated transcription in vitro only from H4 promoters that contained their binding sequences, demonstrating that H4TF-1 and H4TF-2 are H4-specific transcription factors that potentiate expression of this gene.
细胞周期调控的人类组蛋白基因的转录在S期增加约五倍。阐明控制这些基因的细胞周期调控表达的生化机制的第一步是纯化和鉴定调控这些启动子的转录因子。在此，我们描述了两种先前鉴定的因子H4TF-1和H4TF-2的纯化过程，它们可与人组蛋白H4启动子结合。通过离子交换和寡核苷酸亲和层析相结合的方法实现了纯化。基于SDS-聚丙烯酰胺凝胶和紫外线交联对纯化组分的分析，我们认为H4TF-1是由105 kD和110 kD的两种多肽组成。该因子与H4基因最大表达所需的富含GC的DNA序列结合，但不与任何测试的Sp1共有元件结合。H4TF-2是一种65 kD的蛋白质，它特异性地结合到高度保守的H4亚型特异性共有启动子元件内的序列。两种高度纯化的因子仅在含有其结合序列的H4启动子上才能在体外激活转录，这表明H4TF-1和H4TF-2是H4特异性转录因子，可增强该基因的表达。
